KR20200118634A - Novel scyllo inositol-cored amphiphiles and uses thereof - Google Patents

Abstract

The present invention relates to a newly developed scyllo-inositol-based amphiphile compound, a method for preparing the same, and a method for extracting, solubilizing, stabilizing, crystallizing or analyzing a membrane protein by using the same. In addition, this compound can efficiently extract membrane proteins with more various structures and properties from cell membranes than conventional compounds and stably store the membrane proteins in an aqueous solution for a long period of time, and thus can be used for functional analysis and structural analysis accordingly. The analysis of membrane protein structures and functions is one of the most interesting fields in biology and chemistry as it is closely related to the development of novel drugs.

Description

Novel scyllo inositol-cored amphiphiles and uses thereof}

The present invention relates to a newly developed psilo-inositol-based amphiphilic compound, a preparation method thereof, and a method of extracting, solubilizing, stabilizing, crystallizing or analyzing a membrane protein using the same.

Membrain proteins play an important role in biological systems. Since these bio-macromolecules contain hydrophilic and hydrophobic moieties, amphiphilic molecules are required to extract membrane proteins from cell membranes and dissolve and stabilize them in aqueous solutions.

In order to analyze the structure of a membrane protein, a high-quality membrane protein crystal must be obtained, and for this, the structural stability of the membrane protein in an aqueous solution must precede. The number of existing amphiphilic molecules that have been used in membrane protein research is more than 100, and there are many, but only about 5 of them have been actively used in membrane protein structure studies. These five amphoteric molecules are OG (n-octyl-β-D-glucopyranoside), NG (n-nonyl-β-D-glucopyranoside), DM (n-decyl-β-D-maltopyranoside), and DDM (n- dodecyl-β-D-maltopyranoside), and LDAO (lauryldimethylamine- N- oxide) (Non-Patent Document 1, Non-Patent Document 2). However, since many membrane proteins surrounded by these molecules have a tendency to rapidly lose their function due to their structure being easily denatured or aggregated, there are significant limitations in studying the functions and structures of membrane proteins using these molecules. This is because conventional molecules have a simple chemical structure and do not exhibit various properties. Therefore, there is a need to develop a new amphoteric material having new and excellent properties through a new structure.

Accordingly, the present inventors developed an amphiphilic compound in which a hydrophobic group and a hydrophilic group were introduced into the central structure of psilo-inositol, and the present invention was completed by confirming the membrane protein stabilization properties of the compound.

S. Newstead et al., Protein Sci. 17 (2008) 466-472. S. Newstead et al., Mol. Membr. Biol. 25 (2008) 631-638.

An object of the present invention is to provide a compound represented by Formula 1 or Formula 2.

Another object of the present invention is to provide a composition for extraction, solubilization, stabilization, crystallization or analysis of a membrane protein containing the compound.

Another object of the present invention is to provide a method for preparing the compound.

Another object of the present invention is to provide a method for extracting, dissolving, stabilizing, crystallizing or analyzing membrane proteins using the above compound.

One embodiment of the present invention provides a compound represented by the following Formula 1 or Formula 2:

[Formula 1]

In Formula 1,

The R ¹ to R ³ are each independently a substituted or unsubstituted C ₃ -C ₃₀ alkyl group, a substituted or unsubstituted C ₃ -C ₃₀ cycloalkyl group, or a substituted or unsubstituted C ₃ -C ₃₀ aryl Can be a group;

The L ¹ to L ³ may each independently be a direct bond or -O (oxygen)-Y-, wherein Y may be a substituted or unsubstituted C ₁ -C ₁₀ alkylene group; And

The X ¹ to X ³ may be saccharides linked to oxygen.

[Formula 2]

In Chemical Formula 2,

The R ¹ to R ³ are each independently a substituted or unsubstituted C ₃ -C ₃₀ alkyl group, a substituted or unsubstituted C ₃ -C ₃₀ cycloalkyl group, or a substituted or unsubstituted C ₃ -C ₃₀ aryl Can be a group;

The L ⁴ to L ⁶ may be a substituted or unsubstituted C ₁ -C ₁₀ alkylene group; And

The X ⁴ to X ⁹ may be a saccharide linked to oxygen.

As used herein, the term "saccharide" refers to a compound having a relatively small molecule among carbohydrates and having a sweet taste by dissolving in water. Sugars are classified into monosaccharides, disaccharides, and polysaccharides according to the number of molecules constituting the sugar.

The saccharide used in the above embodiments may be monosaccharide or disaccharide, and specifically, may be glucose or maltose, but is not limited thereto.

These saccharides may act as a hydrophilic group. The compound according to an embodiment of the present invention increases the size of the hydrophilic group while minimizing the increase in length by connecting 4 or 8 hydrophilic groups of glucose or maltose in parallel, thereby reducing the size of the complex when forming a complex with a membrane protein. I did. When the size of the complex between the compound and the membrane protein is small, high-quality membrane protein crystals can be obtained (GG Prive, Methods 2007, 41, 388-397).

In addition, R ¹ to R ³ may function as a hydrophobic group. In the compound according to an embodiment of the present invention, three substituted or unsubstituted alkyl groups having different lengths were introduced as hydrophobic groups in order to optimize the balance between hydrophilicity and hydrophobicity.

The compound according to an embodiment of the present invention may have a silo-inositol linker as a central structure. That is, the compound is an amphiphilic material in which 3 or 6 hydrophilic groups and 3 hydrophobic groups are introduced with psilo-inositol as a central structure, and may have excellent performance in stabilizing and crystallizing membrane proteins.

Specifically, R ¹ to R ³ may each independently be a substituted or unsubstituted C ₃ -C ₃₀ alkyl group; In addition, X ¹ to X ⁹ may be glucose or maltose.

In an embodiment of the present invention, in Formula 1, R ¹ to R ³ may each independently be a substituted or unsubstituted C ₃ -C ₃₀ alkyl group; X ¹ to X ³ may be maltose; And the L ¹ to L ³ is a direct bond or -OY-, and Y is such a compound which may be an alkylene group, respectively, "STMs (scyllo-inositol-cored trimaltosides) or STM-Es (scyllo-inositol-cored trimaltosides with ethylene glycol linker)".

In another embodiment of the present invention, in Formula 2, R ¹ to R ³ may each independently be a substituted or unsubstituted C ₃ -C ₃₀ alkyl group; X ⁴ to X ⁹ may be glucose; And the L ⁴ to L ⁶ These compounds, which may be alkylene groups, were each named "SHG-Gs (scyllo-inositol-cored hexaglucoside with glycerol linker)".

In an embodiment of the present invention, as the compound represented by Formula 1, wherein R ¹ to R ³ are C ₁₀ alkyl groups; X ¹ to X ³ are maltose; And L ¹ to L ³ is a direct bond, the compound was named "STM-10". Accordingly, the compound may be a compound represented by Formula 3:

[Formula 3]

In another embodiment of the present invention, as the compound represented by Formula 1, wherein R ¹ to R ³ are C ₁₁ alkyl groups; X ¹ to X ³ are maltose; And L ¹ to L ³ is a direct bond, the compound was named "STM-11". Thus, the compound may be a compound represented by Formula 4:

[Formula 4]

In another embodiment of the present invention, as the compound represented by Formula 1, wherein R ¹ to R ³ are C ₁₂ alkyl groups; X ¹ to X ³ are maltose; And L ¹ to L ³ is a direct bond, the compound was named "STM-12". Thus, the compound may be a compound represented by Formula 5:

[Formula 5]

In another embodiment of the present invention, as the compound represented by Formula 1, wherein R ¹ to R ³ are C ₇ alkyl groups; X ¹ to X ³ are maltose; And L ¹ to L ³ is -O-CH ₂ CH ₂ -The compound was named "STM-E7". Thus, the compound may be a compound represented by Formula 6:

[Formula 6]

In another embodiment of the present invention, as the compound represented by Formula 1, wherein R ¹ to R ³ are C ₈ alkyl groups; X ¹ to X ³ are maltose; And L ¹ to L ³ is -O-CH ₂ CH ₂ -The compound was named "STM-E8". Thus, the compound may be a compound represented by Formula 7:

[Formula 7]

In another embodiment of the present invention, as the compound represented by Formula 1, wherein R ¹ to R ³ are C ₉ alkyl groups; X ¹ to X ³ are maltose; And L ¹ to L ³ is -O-CH ₂ CH ₂ -The compound was named "STM-E9". Thus, the compound may be a compound represented by Formula 8 below:

[Formula 8]

In another embodiment of the present invention, as the compound represented by Formula 1, wherein R ¹ to R ³ are C ₁₀ alkyl groups; X ¹ to X ³ are maltose; And L ¹ to L ³ is -O-CH ₂ CH ₂ -The compound was named "STM-E10". Thus, the compound may be a compound represented by Formula 9:

[Formula 9]

In another embodiment of the present invention, as the compound represented by Formula 2, wherein R ¹ to R ³ are C ₁₂ alkyl groups; X ⁴ to X ⁹ are glucose; And L ⁴ to L ⁶ was named "SHG-G12" a compound of -CH ₂ -. Thus, the compound may be a compound represented by Formula 10 below:

[Formula 10]

In another embodiment of the present invention, as the compound represented by Formula 2, wherein R ¹ to R ³ are C ₁₃ alkyl groups; X ⁴ to X ⁹ are glucose; And L ⁴ to L ⁶ was named "SHG-G13" a compound of -CH ₂ -. Thus, the compound may be a compound represented by the following formula (11):

[Formula 11]

In another embodiment of the present invention, as the compound represented by Formula 2, wherein R ¹ to R ³ are C ₁₄ alkyl groups; X ⁴ to X ⁹ are glucose; And L ⁴ to L ⁶ was named "SHG-G14" a compound of -CH ₂ -. Thus, the compound may be a compound represented by the following Formula 12:

[Formula 12]

The compound according to another embodiment of the present invention may be an amphiphilic molecule for extracting, dissolving, stabilizing, crystallizing or analyzing a membrane protein, but is not limited thereto.

Specifically, the extraction may be to extract the membrane protein from the cell membrane.

As used herein, the term "amphiphilic molecule" refers to a molecule capable of having affinity for both polar and non-polar solvents by coexisting hydrophobic groups and hydrophilic groups in one molecule. Phospholipid molecules present in surfactants or cell membranes have a hydrophilic group at one end and a hydrophobic group at the other end. They have amphiphilic properties and form micelles or liposomes in aqueous solutions. Hydrophilic groups have polarity, but since non-polar groups coexist, their amphiphilic molecules tend to be less soluble in aqueous solutions. However, when the concentration exceeds a certain limit concentration (critical micelle concentration, CMC), hydrophobic groups are collected inside by hydrophobic interaction, and round or elliptical micelles are formed in which the hydrophilic groups are exposed on the surface. .

A method of measuring CMC is not particularly limited, but a method widely known in the art may be used, for example, it may be measured by a fluorescent staining method using diphenylhexatriene (DPH).

The compound according to an embodiment of the present invention may have a critical micelle concentration (CMC) in an aqueous solution of 0.0001 to 1 mM, specifically 0.001 to 0.5 mM, and more specifically 0.005 to 0.3 mM, but is limited thereto. I never do that.

In the case of DDM, which is mainly used for membrane protein studies, most of the STMs, STM-Es or SHG-Gs of this embodiment have a small CMC value compared to the critical micelle concentration of 0.17 mM. Therefore, since micelles are easily formed even at low concentrations of STMs, STM-Es or SHG-Gs, it can be said that it is more advantageous in terms of utilization than DDM because it is possible to study and analyze membrane proteins effectively using a small amount.

In addition, another embodiment of the present invention provides a composition for extraction, solubilization, stabilization, crystallization or analysis of a membrane protein containing the compound.

Specifically, the extraction may be to extract the membrane protein from the cell membrane.

The composition may be in the form of micelles, liposomes, emulsions, or nanoparticles, but is not limited thereto.

The micelles may have a radius of 2.0 nm to 50 nm, and specifically 2.0 nm to 30.0 nm, but are not limited thereto.

A method of measuring the radius of the micelle is not particularly limited, but a method well known in the art may be used, and may be measured using, for example, a dynamic light scattering (DLS) experiment.

The micelles, liposomes, emulsions, or nanoparticles are hydrophobic and can bind to membrane proteins. That is, the micelles, liposomes, emulsions, or nanoparticles may extract and wrap a membrane protein present in a cell membrane. Therefore, it is possible to extract, solubilize, stabilize, crystallize, or analyze membrane proteins from the cell membrane by the micelles.

The composition may further include a buffer that may aid in extraction, solubilization, stabilization, crystallization, or analysis of membrane proteins.

In addition, another embodiment of the present invention provides a method for preparing a compound represented by the following formula (1) comprising the steps of 1) to 4) below:

1) performing an alkylation reaction by adding alkyl iodide to silitol-1, 3, 5-orthoacetate;

2) treating the product of step 1) with an acid to remove the orthoacetate protecting group to produce a triol;

3) introducing a saccharide having a protecting group attached thereto by performing a glycosylation reaction to the product of step 2); And

4) performing a deprotection reaction on the product of step 3),

[Formula 1]

In Formula 1,

The R ¹ to R ³ are each independently a substituted or unsubstituted C ₃ -C ₃₀ alkyl group, a substituted or unsubstituted C ₃ -C ₃₀ cycloalkyl group, or a substituted or unsubstituted C ₃ -C ₃₀ aryl Can be a group;

The L ¹ to L ³ may be a direct bond; And

The X ¹ to X ³ may be saccharides linked to oxygen.

The compound synthesized by the above method may be one of Chemical Formulas 3 to 5 according to an embodiment of the present invention, but is not limited thereto.

In this embodiment, since the compound can be synthesized by a simple method through five synthesis steps, mass production of the compound for membrane protein research is possible.

In addition, another embodiment of the present invention provides a method for preparing a compound represented by the following formula 1, including the steps of 1) to 5):

1) performing an alkylation reaction by adding an alkyl iodide to 1,3,5-triallylated scyllitol;

2) performing an ozonolysis reaction on the product of step 1);

3) introducing a saccharide to which a protecting group is attached by performing a glycosylation reaction to the product of step 2; And

4) performing a deprotection reaction on the product of step 3),

[Formula 1]

In Formula 1,

The R ¹ to R ³ are each independently a substituted or unsubstituted C ₃ -C ₃₀ alkyl group, a substituted or unsubstituted C ₃ -C ₃₀ cycloalkyl group, or a substituted or unsubstituted C ₃ -C ₃₀ aryl Can be a group;

The L ¹ to L ³ may be -O (oxygen)-Y-, and Y may be a substituted or unsubstituted C ₁ -C ₁₀ alkylene group; And

The X ¹ to X ³ may be saccharides linked to oxygen.

The compound synthesized by the above method may be one of Chemical Formulas 6 to 9 according to an embodiment of the present invention, but is not limited thereto.

In addition, another embodiment of the present invention provides a method for preparing a compound represented by the following formula (2) comprising the steps of 1) to 5) below:

1) performing an alkylation reaction by adding an alkyl iodide to 1,3,5-triallylated scyllitol;

2) performing dihydroxylation on the product of step 1);

3) introducing a saccharide to which a protecting group is attached by performing a glycosylation reaction to the product of step 2; And

4) performing a deprotection reaction on the product of step 3),

[Formula 2]

In Chemical Formula 2,

The R ¹ to R ³ are each independently a substituted or unsubstituted C ₃ -C ₃₀ alkyl group, a substituted or unsubstituted C ₃ -C ₃₀ cycloalkyl group, or a substituted or unsubstituted C ₃ -C ₃₀ aryl Can be a group;

The L ⁴ to L ⁶ may be a substituted or unsubstituted C ₁ -C ₁₀ alkylene group; And

The X ⁴ to X ⁹ may be a saccharide linked to oxygen.

The compound synthesized by the above method may be one of Formulas 10 to 12 according to an embodiment of the present invention, but is not limited thereto.

Another embodiment of the present invention provides a method for extracting, dissolving, stabilizing, crystallizing or analyzing membrane proteins. Specifically, it provides a method for extracting, dissolving, stabilizing, crystallizing or analyzing a membrane protein, comprising the step of treating a membrane protein with a compound represented by the following Formula 1 or Formula 2 in an aqueous solution:

[Formula 1]

In Formula 1,

The R ¹ to R ³ are each independently a substituted or unsubstituted C ₃ -C ₃₀ alkyl group, a substituted or unsubstituted C ₃ -C ₃₀ cycloalkyl group, or a substituted or unsubstituted C ₃ -C ₃₀ aryl Can be a group;

The L ¹ to L ³ may each independently be a direct bond or -O (oxygen)-Y-, wherein Y may be a substituted or unsubstituted C ₁ -C ₁₀ alkylene group; And

The X ¹ to X ³ may be saccharides linked to oxygen.

[Formula 2]

In Chemical Formula 2,

The R ¹ to R ³ are each independently a substituted or unsubstituted C ₃ -C ₃₀ alkyl group, a substituted or unsubstituted C ₃ -C ₃₀ cycloalkyl group, or a substituted or unsubstituted C ₃ -C ₃₀ aryl Can be a group;

The L ⁴ to L ⁶ may be a substituted or unsubstituted C ₁ -C ₁₀ alkylene group; And

The X ⁴ to X ⁹ may be a saccharide linked to oxygen.

Specifically, in Formula 1, R ¹ to R ³ may each independently be a substituted or unsubstituted C ₃ -C ₃₀ alkyl group; X ¹ to X ³ may be maltose; And the L ¹ to L ³ is a direct bond or -OY-, and the Y may be an alkylene group.

In another embodiment of the present invention, in Formula 2, R ¹ to R ³ may each independently be a substituted or unsubstituted C ₃ -C ₃₀ alkyl group; X ⁴ to X ⁹ may be glucose; And the L ⁴ to L ⁶ may be an alkylene group.

The compound may be one of Formulas 3 to 12 according to an embodiment of the present invention, but is not limited thereto.

Specifically, the extraction may be to extract the membrane protein from the cell membrane.

As used herein, the term "membrane protein" is a generic term for a protein or glycoprotein that is transferred to the cell membrane lipid bilayer. These exist in various states, such as penetrating the entire cell membrane, being located on the surface, or adhering to the cell membrane. Examples of membrane proteins include, but are not limited to, enzymes, receptors such as peptide hormones and local hormones, receptor carriers such as sugars, ion channels, and cell membrane antigens.

The membrane protein includes any protein or glycoprotein that is transferred to the cell membrane lipid bilayer, specifically LHI-RC, LeuT (Leucine transporter), β ₂ AR (human β ₂ adrenergic receptor), MelB _st (melibiose permease) or a combination of two or more thereof, but is not limited thereto.

As used herein, the term "extraction of membrane proteins" means separating membrane proteins from cell membranes.

As used herein, the term "solubilization of membrane proteins" refers to dissolving membrane proteins insoluble in water into micelles in an aqueous solution.

As used herein, the term "stabilization of a membrane protein" means stably preserving a tertiary or quaternary structure so that the structure and function of the membrane protein are not changed.

As used herein, the term "crystallization of a membrane protein" means forming a crystal of a membrane protein in a solution.

As used herein, the term "analysis of a membrane protein" means analyzing the structure or function of a membrane protein. In the above embodiment, the analysis of the membrane protein may use a known method, but is not limited thereto, for example, analyzing the structure of the membrane protein using electron microscopy or nuclear magnetic resonance. can do.

When the silo-inositol-based compound according to the embodiments of the present invention is used, the membrane protein can be stably stored in an aqueous solution for a long time compared to the existing compound, and through this, it can be used for functional analysis and structural analysis.

Membrane protein structure and function analysis is one of the fields of interest in current biology and chemistry, so it can be applied to protein structure studies that are closely related to the development of new drugs.

In addition, since the compounds according to the embodiments of the present invention can be synthesized by a simple method from readily available starting materials, mass production of compounds for membrane protein studies is possible.

1 is a diagram showing a synthesis scheme of STMs according to Example 1 of the present invention.
2 is a diagram showing a synthesis scheme of STM-Es according to Example 2 of the present invention.
3 is a diagram showing a synthesis scheme of SHG-Gs according to Example 3 of the present invention.
4 shows the DLS profile of STMs, STM-Es or SHG-Gs.
5 is a diagram showing the results of measuring the long-term stability of R. capsulatus assembly containing LHI-RC dissolved by CMC + 0.2 wt% STMs, STM-Es or SHG-Gs.
6 is a result of measuring the structural stability of LeuT (Leucine transporter) in an aqueous solution by CMC + 0.2 wt% of STMs, STM-Es, SHG-Gs or DDM. Protein stability was confirmed by measuring the substrate binding properties of the transporter through a scintillation proximity assay (SPA). In the presence of each amphiphilic compound, LeuT was incubated at room temperature for 14 days to measure the protein's substrate binding properties at regular intervals:
Figure 7 is a MelB _St protein using STMs, STM-Es, SHG-Gs or DDM at a concentration of 1.5 wt%, extracted at four temperatures (0, 45, 55, 65 ℃), incubated at the same temperature for 90 minutes Then, the amount of MelB _St protein dissolved in the aqueous solution was measured:
(a) SDS-PAGE and Western Blotting results showing the amount of MelB protein extracted using each amphiphilic compound, and the amount of MelB _St protein extracted using each amphiphilic compound is present in a membrane sample (Memb) without an amphiphilic compound A histogram expressed as a percentage (%) of the total amount of protein to be taken; And
(b) As a result of checking whether the function of MelB dissolved by DDM or STM-12 is maintained
Figure 8 shows the effect on the stability of β ₂ AR by STMs, STM-Es, SHG-Gs or DDM (a) at 30 minutes, (b) at regular intervals for 3 days, or (c) at regular intervals for 5 days It is the result of measurement. Protein ligand binding properties were measured through a ligand binding assay of [ ³ H]-dihydroalprenolol (DHA).

Hereinafter, the present invention will be described in more detail in the following examples. However, the following examples are not intended to limit or limit the scope of the present invention only to illustrate the content of the present invention. What can be easily inferred by those skilled in the art from the detailed description and examples of the present invention is construed as belonging to the scope of the present invention.

<Example 1> Synthesis method of STMs

The synthesis scheme of STMs is shown in FIG. 1. Three kinds of STMs compounds were synthesized according to the synthesis methods of the following <1-1> to <1-3>.

<1-1> 1,3,5- Trialkylated Silitol (1,3,5- trialkylated scyllitol Synthesis of (Compound A) (Step a in Fig. 1)

After cooling the solution of silitol-1,3,5-orthoacetate (compound 1a, 1.0 equivalent) mixed in DMF to 0 °C, NaH (mineral oil) was added under an inert atmosphere. 60% dispersion, 6.0 eq) was added. Alkyl iodide (4.5 equivalents) was added dropwise after gas evolution ceased, and the reaction mixture was stirred at room temperature until thoroughly mixed. After adding ice, it was diluted with distilled water to quench the reaction. The diluted reaction mixture was extracted with ethyl acetate. The combined ethyl acetate fractions were washed with brine and dried over anhydrous Na ₂ SO ₄ . Removal of the solvent under reduced pressure gave a colorless oil dissolved in a TFA/H2O (10:1) mixture, and stirring was continued at room temperature for 2 hours. TFA was rotary evaporated with co-solvent DCM to give an oily residue, which was redissolved in DCM/MeOH (1: 1). NaOH (25% by weight) was added to the mixture to increase the pH by 12 or more. The resulting alkaline mixture was stirred at 40° C. for 4 hours and then diluted with water. The diluted mixture was washed twice with DCM. The combined organic layer was washed with brine and dried over anhydrous Na ₂ SO ₄ . DCM was removed under reduced pressure to obtain an oily residue, and the title compound A was obtained as a colorless oil through column chromatography purification.

<1-2> General synthesis procedure of maltosylation reaction (step b of FIG. 1)

Under nitrogen (N ₂ ), a mixture of compound A (1.0 equivalent), dimethoxyethane (30.0 equivalent) and AgOTf (3.6 equivalent) was stirred in anhydrous CH ₂ Cl ₂ at -45°C. A solution of perbenzoylated maltosylbromide (3.6 equivalents) in anhydrous CH ₂ Cl ₂ was added dropwise to this suspension. After continuing the stirring at -45° C. for 5 minutes, the temperature of the reaction mixture was increased to 0° C. and stirring was continued for 30 minutes. After completion of the reaction (represented by TLC), pyridine was added to quench the reaction, then diluted with CH ₂ Cl ₂ and filtered over Celite. The filtrate was washed successively with 1 M aqueous Na ₂ S ₂ O ₃ solution, 0.1 M HCl aqueous solution, and brine. After drying the organic layer with anhydrous Na ₂ SO ₄ , the solvent was removed under reduced pressure. The obtained residue was purified by silica gel column chromatography (EtOAc/hexane) to obtain the desired product compound A ₁ as a glassy solid.

<1-3> Deprotection vaporization reaction ( deprotection reaction) for general synthesis procedure (step c in FIG. 1)

This was followed by the synthesis method of Chae, PS et al. (Nat Meth 2010, 7, 1003.). De-O-benzoylation was performed under Zemplen's conditions. The O-protected compound was dissolved in anhydrous CH2Cl2 and then MeOH was slowly added until a continuous precipitation appeared. NaOMe, which is a 0.5M methanolic solution, was added to the reaction mixture so that the final concentration was 0.05M. The reaction mixture was stirred at room temperature for 6 hours. After completion of the reaction, the reaction mixture was neutralized using Amberlite IR-120 (H+ form) resin. The resin was removed by filtration, washed with MeOH, and the solvent was removed from the filtrate under vacuum conditions (in vacuo). Diethyl ether (50 mL) was added to the residue dissolved in 2 mL of MeOH:CH ₂ Cl ₂ (1:1) mixture to obtain the target compound A ₂ as a precipitated white solid.

<Production Example 1> Synthesis of STM-10

<1-1> Synthesis of Compound A1

Compound A1 was synthesized in 70% yield according to the procedure for synthesizing 1,3,5-trialkylated silitol of Example 1-1. ¹ H NMR (400 MHz, CDCl ₃ ): δ 3.79 (t, J = 6.0 Hz, 6H), 3.44 (t, J = 10.0 Hz, 3H), 3.11 (t, J = 10.0 Hz, 3H), 3.03 ( s br, 3H), 1.60 (quin, J = 6.0 Hz, 6H), 1.38-1.20 (m, 42H), 0.88 (t, J = 8.0 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ): δ 82.3, 74.2, 32.1, 30.5, 29.8, 29.7, 29.6, 29.5, 26.3, 22.8, 14.3.

<1-2> Synthesis of STM-10a

Compound STM-10a was synthesized in a yield of 62% according to the general saccharification reaction procedure of Example 1-2. ¹ H NMR (400 MHz, CDCl ₃ ): δ 8.14 (d, J = 8.0 Hz, 6H), 7.97 (d, J = 8.0 Hz, 6H), 7.83 (d, J = 8.0 Hz, 6H), 7.73- 7.68 (m, 18H), 7.59 (t, J = 10.0 Hz, 3H), 7.54 (d, J = 8.0 Hz, 6H), 7.52-7.45 (m, 12H), 7.43-7.38 (m, 12H), 7.36 -7.30 (m, 12H), 7.25-7.21 (m, 11H), 7.20 (d, J = 8.0 Hz, 6H), 7.13 (t, J = 8.0 Hz, 7H), 6.14 (t, J = 10.0 Hz, 3H), 5.80-5.66 (m, 9H), 5.35-5.30 (m, 8H), 5.23 (d, J = 4.0 Hz, 3H), 5.19 (t, J = 10.0 Hz, 3H), 4.92 (d, J = 12.0 Hz, 3H), 4.81 (d, J = 8.0 Hz, 3H), 4.49 (t, J = 8.0 Hz, 3H), 4.40-4.37 (m, 3H), 4.33-4.30 (m, 3H), 4.18 (dd, J _1-2 = 4.0 Hz, J _1-3 = 12.0 Hz, 3H), 4.04 (d, J = 12.0 Hz, 3H), 3.66-3.57 (m, 6H), 3.12 (q, J = 8.0 Hz, 3H), 2.85 (t, J = 10.0 Hz, 3H), 1.48-1.38 (m, 6H), 1.28-1.08 (m, 42H), 0.88 (t, J = 6.0 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ): δ 166.3, 165.8, 165.6, 165.2, 164.9, 133.7, 133.5, 133.3, 133.1, 130.1, 130.0, 129.9, 129.8, 129.7, 129.5, 129.2, 129.1, 128.9, 128.8, 128.6, 128.5, 128.3, 128.2, 100.2, 96.4, 80.8, 78.6, 75.2, 73.5, 72.8, 71.0, 70.0, 69.2, 63.9, 62.6, 32.2, 30.3, 30.0, 29.8, 29.7, 26.3, 23.0, 14.4.

<1-3> Synthesis of STM-10

According to the general synthetic procedure for the deprotection vaporization reaction of Example 1-3, STM-10 was obtained in a yield of 91%. Synthesized. ¹ H NMR (400 MHz, CD ₃ OD): δ 5.11 (d, J = 4.0 Hz, 3H), 4.79 (d, J = 8.0 Hz, 3H), 3.94-3.90 (m, 6H), 3.82-3.77 ( m, 9H), 3.71-3.64 (m, 9H), 3.62-3.58 (m, 6H), 3.44 (dd, J _1-2 = 4.0 Hz, J _1-3 = 12.0 Hz, 3H), 3.39 (t, J = 8.0 Hz, 3H), 3.34-3.27 (m, 9H), 3.24-3.19 (dd, J _1-2 = 4.0 Hz, J _1-3 = 8.0 Hz, 3H), 1.64-1.62 (m, 6H) , 1.39-1.21 (m, 42H), 0.90 (t, J = 8.0 Hz, 9H); ¹³ C NMR (100 MHz, CD ₃ OD): δ 105.0, 102.9, 81.3, 77.9, 76.5, 75.6, 75.1, 74.8, 74.2, 71.3, 70.4, 70.1, 62.7, 62.3, 48.5, 46.6, 41.6, 39.5, 32.7 , 27.7, 23.8, 14.7. HRMS ( FAB ⁺ ) : calcd. for C ₇₂ H ₁₃₂ O ₃₆ [M+Na] ⁺ 1595.8396, found 1595.8402.

<Production Example 2> Synthesis of STM-11

<2-1> Synthesis of Compound A2

Compound A2 was synthesized in 70% yield according to the procedure for synthesizing 1,3,5-trialkylated silitol of Example 1-1. ¹ H NMR (400 MHz, CDCl ₃ ): δ 3.79 (t, J = 6.0 Hz, 6H), 3.43 (t, J = 8.0 Hz, 3H), 3.10 (t, J = 10.0 Hz, 3H), 2.69 ( s br, 3H), 1.60 (quin, J = 8.0 Hz, 6H), 1.38-1.20 (m, 48H), 0.88 (t, J = 8.0 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ): δ 82.3, 74.2, 73.5, 32.1, 30.5, 29.8, 29.7, 29.6, 29.5, 26.3, 22.8, 14.3.

<2-2> Synthesis of STM-11a

Compound STM-11a was synthesized in a yield of 63% according to the general saccharification reaction procedure of Example 1-2. ¹ H NMR (400 MHz, CDCl ₃ ): δ 8.15 (d, J = 8.0 Hz, 6H), 7.99 (d, J = 8.0 Hz, 6H), 7.80 (d, J = 8.0 Hz, 6H), 7.73- 7.70 (m, 18H), 7.59 (t, J = 10.0 Hz, 3H), 7.54 (d, J = 8.0 Hz, 6H), 7.52-7.45 (m, 12H), 7.43-7.38 (m, 12H), 7.36 -7.30 (m, 12H), 7.25-7.21 (m, 11H), 7.20 (d, J = 8.0 Hz, 6H), 7.13 (t, J = 8.0 Hz, 7H), 6.06 (t, J = 10.0 Hz, 3H), 5.72 (t, J = 10.0 Hz, 3H), 5.67 (d, J = 4.0 Hz, 3H), 5.63 (t, J = 8.0 Hz, 3H), 5.28 (t, J = 10.0 Hz, 3H) , 5.23 (d, J = 4.0 Hz, 3H), 5.19 (t, J = 10.0 Hz, 3H), 4.92 (d, J = 12.0 Hz, 3H), 4.81 (d, J = 8.0 Hz, 3H), 4.49 (t, J = 8.0 Hz, 3H), 4.40-4.37 (m, 3H), 4.33-4.30 (m, 3H), 4.18 (dd, J _1-2 = 4.0 Hz, J _1-3 = 12.0 Hz, 3H ), 4.04 (d, J = 12.0 Hz, 3H), 3.66-3.57 (m, 6H), 3.12 (q, J = 8.0 Hz, 3H), 2.85 (t, J = 10.0 Hz, 3H), 1.48-1.38 (m, 6H), 1.28-1.08 (m, 48H), 0.87 (t, J = 8.0 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ): δ 166.3, 165.9, 165.6, 165.3, 165.2, 164.9, 133.7, 133.5, 133.3, 133.1, 130.1, 130.0, 129.9, 129.8, 129.7, 129.6, 129.2, 129.1, 128.9, 128.8, 128.6, 128.5, 128.3, 128.2, 100.1, 96.4, 80.8, 78.6, 75.2, 73.5, 72.8, 72.6, 71.0, 70.0, 69.2, 63.9, 62.6, 32.2, 30.3, 30.0, 29.8, 29.7, 26.3, 23.0, 14.4.

<2-3> Synthesis of STM-11

According to the general synthetic procedure for the deprotection vaporization reaction of Example 1-3, STM-11 was obtained in a yield of 92%. Synthesized. ¹ H NMR (400 MHz, CD ₃ OD): δ 5.11 (d, J = 4.0 Hz, 3H), 4.79 (d, J = 8.0 Hz, 3H), 3.96-3.90 (m, 6H), 3.86-3.77 ( m, 9H), 3.71-3.64 (m, 9H), 3.62-3.58 (m, 6H), 3.44 (dd, J _1-2 = 4.0 Hz, J _1-3 = 12.0 Hz, 3H), 3.39 (t, J = 8.0 Hz, 3H), 3.34-3.27 (m, 9H), 3.24-3.19 (dd, J _1-2 = 8.0 Hz, J _1-3 = 12.0 Hz, 3H), 1.64-1.62 (m, 6H) , 1.39-1.21 (m, 48H), 0.90 (t, J = 8.0 Hz, 9H); ¹³ C NMR (100 MHz, CD ₃ OD): δ 105.0, 102.9, 81.3, 77.8, 76.5, 75.5, 75.1, 74.8, 74.2, 71.5, 70.4, 70.1, 62.7, 62.3, 48.5, 46.6, 41.6, 39.5, 33.6 , 32.7, 27.7, 23.8, 14.7. HRMS ( FAB ⁺ ) : calcd. for C ₇₅ H ₁₃₈ O ₃₆ [M+Na] ⁺ 1637.8866, found 1637.8862.

<Production Example 3> Synthesis of STM-12

<3-1> Synthesis of Compound A3

Compound A3 was synthesized in 72% yield according to the procedure for synthesizing 1,3,5-trialkylated silitol of Example 1-1. ¹ H NMR (400 MHz, CDCl ₃ ): δ 3.91 (t, J = 6.0 Hz, 6H), 3.77 (t, J = 6.0 Hz, 3H), 3.71 (t, J = 8.0 Hz, 3H), 3.04 ( s br, 3H), 1.60 (quin, J = 8.0 Hz, 6H), 1.38-1.20 (m, 54H), 0.88 (t, J = 8.0 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ): δ 82.4, 74.1, 73.4, 63.1, 32.9, 32.1, 30.5, 29.8, 29.7, 29.6, 29.5, 26.3, 25.9, 22.9, 14.3.

<3-2> Synthesis of STM-12a

Compound STM-12a was synthesized in a yield of 62% according to the general saccharification reaction procedure of Example 1-2. ¹ H NMR (400 MHz, CDCl ₃ ): δ 8.15 (d, J = 8.0 Hz, 6H), 8.00 (d, J = 8.0 Hz, 6H), 7.80 (d, J = 8.0 Hz, 6H), 7.73- 7.69 (m, 18H), 7.61 (t, J = 6.0 Hz, 3H), 7.55-7.52 (m, 6H), 7.48-7.44 (m, 12H), 7.42-7.38 (m, 12H), 7.36-7.31 ( m, 12H), 7.26-7.19 (m, 18H), 7.16 (t, J = 8.0 Hz, 6H), 6.05 (t, J = 10.0 Hz, 3H), 5.71 (t, J = 10.0 Hz, 3H), 5.67 (d, J = 4.0 Hz, 3H), 5.63 (t, J = 8.0 Hz, 3H), 5.27 (t, J = 8.0 Hz, 3H), 5.23 (d, J = 8.0 Hz, 3H) 5.19 (t , J = 8.0 Hz, 3H), 4.91 (d, J = 8.0 Hz, 3H), 4.81 (d, J = 12.0 Hz, 3H), 4.49 (t, J = 10.0 Hz, 3H), 4.40-4.36 (m , 3H), 4.33-4.29 (m, 3H), 4.18 (dd, J _1-2 = 4.0 Hz, J _1-3 = 12.0 Hz, 3H), 4.04 (d, J = 8.0 Hz, 3H), 3.65- 3.57 (m, 6H), 3.11 (q, J = 8.0 Hz, 3H), 2.85 (t, J = 10.0 Hz, 3H), 1.48-1.38 (m, 6H), 1.28-1.08 (m, 54H), 0.87 (t, J = 8.0 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ): δ 166.2, 165.8, 165.5, 165.2, 165.1, 164.8, 133.6, 133.4, 133.2, 133.1, 130.0, 129.9, 129.7, 129.6, 129.5, 129.4, 129.1, 128.9, 128.8, 128.6, 128.5, 128.4, 128.2, 100.0, 96.3, 80.7, 78.5, 75.1, 73.4, 72.4, 70.9, 69.8, 69.1, 63.9, 62.5, 53.5, 32.1, 30.2, 29.9, 29.8, 29.7, 29.5, 26.2, 22.8, 21.1, 14.3.

<3-3> Synthesis of STM-12

According to the general synthetic procedure for the deprotection vaporization reaction of Example 1-3, STM-12 in a yield of 92% Synthesized. ¹ H NMR (400 MHz, CD ₃ OD): δ 5.14 (d, J = 4.0 Hz, 3H), 4.27 (d, J = 8.0 Hz, 3H), 3.99 (q, J = 8.0 Hz, 3H), 3.86 -3.71 (m, 15H), 3.69-3.59 (m, 24H), 3.53 (t, J = 8.0 Hz, 3H), 3.44 (dd, J _1-2 = 4.0 Hz, J _1-3 = 8.0 Hz, 3H ), 3.36-3.31 (m, 3H), 3.28-3.22 (m, 3H), 1.59-1.57 (m, 6H), 1.39-1.21 (m, 54H), 0.90 (t, J = 8.0 Hz, 9H); ¹³ C NMR (100 MHz, CD ₃ OD): δ 104.5, 103.1, 84.1, 81.5, 77.9, 76.8, 75.2, 75.0, 74.9, 74.8, 74.3, 71.7, 71.6, 68.4, 62.8, 62.3, 33.2, 32.1, 31.8 , 31.0, 30.9, 30.6, 27.6, 23.9, 14.7. HRMS (FAB ⁺ ) : calcd. for C ₇₈ H ₁₄₄ O ₃₆ [M+Na] ⁺ 1679.9335, found 1679.9341.

<Example 2> Synthesis method of STM-Es

The synthesis scheme of STM-Es is shown in FIG. 2. According to the synthesis method of the following <2-1> to <2-4>, three types of STM-Es compounds were synthesized.

<2-1> 1,3,5- Triallylation -2,4,6- Trialkylated Silitol (1,3,5-triallylated-2,4,6-trialkylated scyllitol ) Synthesis of (Compound B) (Step a in Fig. 2)

NaH (60% dispersion in mineral oil, 6.0 equivalents) was added to 1,3,5-triallylated syllitol (1,3,5-triallylated syllitol) (compound 1b) dissolved in DMF under argon (Ar) atmosphere. I did. Alkyl iodide (4.5 equivalents) was added dropwise after gas evolution ceased, and the reaction mixture was stirred at room temperature for 24 hours and ice was added to quench the reaction. The quenched reaction mixture was diluted with water and then washed with ethyl acetate. The combined ethyl acetate fractions were washed with brine and dried over anhydrous Na ₂ SO ₄ . The solvent was removed under reduced pressure to obtain an oily residue, and purified by column chromatography to give the title compound B as a colored oil.

<2-2> General synthesis procedure of ozonolysis reaction (step b in Fig. 2)

A mixture of 1,3,5-triallylated-2,4,6-trialkylated silitol dissolved in MeOH was cooled to -78°C. Ozone was purged into the reaction mixture for 15 minutes until a blue color was maintained. After the reaction was stirred at -78 °C for 20 minutes, dimethyl sulfide (8.0 equivalents) was added. The temperature was slowly increased and the mixture was stirred at room temperature for 18 hours. The organic solvent was removed from the reaction mixture to give an oily residue, which was purified by column chromatography to give the title compound C as a colorless oil.

<2-3> General synthesis procedure of maltosylation reaction (step c in FIG. 2)

Under nitrogen (N ₂ ), a mixture of compound A (1.0 equivalent), dimethoxyethane (30.0 equivalent) and AgOTf (3.6 equivalent) was stirred in anhydrous CH ₂ Cl ₂ at -45°C. A solution of perbenzoylated maltosylbromide (3.6 equivalents) in anhydrous CH ₂ Cl ₂ was added dropwise to this suspension. After continuing the stirring at -45° C. for 5 minutes, the temperature of the reaction mixture was increased to 0° C. and stirring was continued for 30 minutes. After completion of the reaction (represented by TLC), pyridine was added to quench the reaction, then diluted with CH ₂ Cl ₂ and filtered over Celite. The filtrate was washed successively with 1 M aqueous Na ₂ S ₂ O ₃ solution, 0.1 M HCl aqueous solution, and brine. After drying the organic layer with anhydrous Na ₂ SO ₄ , the solvent was removed under reduced pressure. The obtained residue was purified by silica gel column chromatography (EtOAc/hexane) to obtain the desired product compound C ₁ as a glassy solid.

<2-4> Deprotection vaporization reaction ( deprotection reaction) for general synthesis procedure (step d in FIG. 2)

This was followed by the synthesis method of Chae, PS et al. (Nat Meth 2010, 7, 1003.). De-O-benzoylation was performed under Zemplen's conditions. The O-protected compound was dissolved in anhydrous CH2Cl2 and then MeOH was slowly added until a continuous precipitation appeared. NaOMe, which is a 0.5M methanolic solution, was added to the reaction mixture so that the final concentration was 0.05M. The reaction mixture was stirred at room temperature for 6 hours. After completion of the reaction, the reaction mixture was neutralized using Amberlite IR-120 (H+ form) resin. The resin was removed by filtration, washed with MeOH, and the solvent was removed from the filtrate under vacuum conditions (in vacuo). Diethyl ether (50 mL) was added to the residue dissolved in 2 mL of MeOH:CH ₂ Cl ₂ (1:1) mixture to obtain the target compound C ₂ as a precipitated white solid.

<Production Example 4> Synthesis of STM-E7

<4-1> Synthesis of Compound B4

Compound B4 was synthesized in 75% yield according to the general alkylation reaction procedure of 1,3,5-triallylated silitol of Example 2-1. ¹ H NMR (400 MHz, CDCl ₃ ): δ 5.98-5.91 (m, 3H), 5.26 (dd, J _1-2 = 4.0 Hz, J _1-3 = 20.0 Hz, 3H), 5.14 (dd, J _{1 -2} = 4.0 Hz, J _{1 -3} = 12.0 Hz, 3H), 4.27 (d, J = 8.0 Hz, 6H), 3.73 (t, J = 6.0 Hz, 6H), 3.10 (s, 6H), 1.58 ( quin, J = 8.0 Hz, 6H), 1.31-1.20 (m, 24H), 0.88 (t, J = 6.0 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ): δ 135.2, 116.5, 82.7, 82.4, 77.4, 77.1, 76.7, 74.5, 74.0, 31.8, 30.5, 29.2, 26.1, 23.0, 14.0.

<4-2> Synthesis of Compound C8

Compound C8 was synthesized with 71% yield according to the general ozone decomposition reaction procedure of Example 2-2. ¹ H NMR (400 MHz, CDCl ₃ ): δ 3.88-3.86 (m, 6H), 3.78 (t, J = 6.0 Hz, 6H), 3.70 (t, J = 6.0 Hz, 6H), 3.45 (br s, 3H), 3.28 (t, J = 10.0 Hz, 3H), 3.10 (t, J = 10.0 Hz, 3H) , 1.64 (quin, J = 6.0 Hz, 6H), 1.31-1.27 (m, 24H), 0.88 (t, J = 6.0 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ): δ 83.0, 82.9, 74.8, 74.7, 68.2, 31.9, 30.3, 29.3, 26.1, 22.7, 14.2.

<4-3> Synthesis of STM-E7a

Compound STM-E7a was synthesized in a yield of 65% according to the general saccharification reaction procedure of Example 2-3. ¹ H NMR (400 MHz, CDCl ₃ ): δ 8.10 (d, J = 8.0 Hz, 6H), 7.98 (d, J = 8.0 Hz, 6H), 7.85 (d, J = 4.0 Hz, 6H), 7.77 ( d, J = 8.0 Hz, 6H), 7.74 (dd, J _1-2 = 4.0 Hz, J _1-3 = 12.0 Hz, 12H), 7.63 (d, J = 8.0 Hz, 6H), 7.51-7.48 (m , 6H), 7.46-7.35 (m, 24H), 7.31-7.29 (m, 9H), 7.27-7.16 (m, 24H), 6.08 (t, J = 8.0 Hz, 3H), 5.76-5.72 (m, 6H) ), 5.65 (t, J = 10.0 Hz, 3H), 5.30 (t, J = 10.0 Hz, 6H), 5.25 (dd, J _1-2 = 4.0 Hz, J _1-3 = 12.0 Hz, 3H), 4.90 (d, J = 12.0 Hz, 3H), 4.80 (d, J = 4.0 Hz, 3H), 4.74 (dd, J _1-2 = 4.0 Hz, J _1-3 = 12.0 Hz, 3H), 4.49 (t, J = 8.0 Hz, 3H), 4.46-4.40 (m, 3H), 4.38 (dd, J _1-2 = 4.0 Hz, J _1-3 = 12.0 Hz, 3H), 4.24 (dd, J _1-2 = 4.0 Hz, J _1-3 = 12.0 Hz, 3H), 4.08 (d, J = 12.0 Hz, 3H), 3.93-3.90 (m, 3H), 3.72-3.70 (m, 3H), 3.63-3.57 (m, 6H) ), 3.42 (q, J = 4.0 Hz, 3H), 3.33 (q, J = 4.0 Hz, 3H), 2.64 (t, J = 12.0 Hz, 3H), 2.42 (t, J = 8.0 Hz, 3H) 1.32 -1.30 (m, 6H), 1.25-1.08 (m, 24H), 0.86 (t, J = 6.0 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ): δ 166.3, 165.9, 165.8, 165.6, 165.2, 165.1, 133.6, 133.5, 133.4, 133.3, 133.2, 130.1, 130.0, 129.9, 129.7, 129.6, 129.5, 129.3, 129.0, 128.9, 128.7, 128.6, 128.5, 128.4, 128.3, 128.2, 101.1, 96.5, 83.1, 82.0, 75.3, 73.6, 73.3, 73.1, 72.3, 71.6, 71.0, 70.0, 69.4, 69.2, 63.7, 53.6, 32.0, 30.5, 29.4, 26.4, 22.9, 14.3.

<4-4> Synthesis of STM-E7

According to the general synthetic procedure for the deprotection vaporization reaction of Example 2-4, STM-E7 in a yield of 92% Synthesized. ¹ H NMR (400 MHz, CD ₃ OD): δ 5.15 (d, J = 4.0 Hz, 3H), 4.32 (d, J = 8.0 Hz, 3H), 4.05-3.95 (m, 6H), 3.90 (d, J = 12.0 Hz, 6H), 3.86 (d, J = 16.0 Hz, 6H), 3.82-3.71 (m, 15H), 3.69-3.60 (m, 12H), 3.53 (t, J = 10.0 Hz, 3H), 3.44 (dd, J _1-2 = 4.0 Hz, J _1-3 = 12.0 Hz, 3H), 3.39-3.36 (m, 3H), 3.2 (t, J = 8.0 Hz, 3H), 3.09 (t, J = 8.0 Hz, 3H), 1.64-1.62 (m, 6H), 1.39-1.21 (m, 24H), 0.91 (t, J = 6.0 Hz, 9H); ¹³ C NMR (100 MHz, CD ₃ OD): δ 104.3, 103.3, 84.3, 83.8, 81.4, 77.8, 76.7, 75.1, 74.8, 74.2, 73.4, 71.5, 70.4, 62.8, 62.3, 33.1, 31.7, 30.6, 27.4 , 23.8, 14.7. HRMS ( FAB ⁺ ) : calcd. for C ₆₉ H ₁₂₆ O ₃₉ [M+Na] ⁺ 1601.7774, found 1601.7771.

<Production Example 5> Synthesis of STM-E8

<5-1> Synthesis of Compound B5

Compound B5 was synthesized in 73% yield according to the general alkylation reaction procedure of 1,3,5-triallylated silitol of Example 2-1. ¹ H NMR (400 MHz, CDCl ₃ ): δ 5.99-5.91 (m, 3H), 5.26 (dd, J _1-2 = 3.0 Hz, J _1-3 = 16.0 Hz, 3H), 5.14 (dd, J _{1 -2} = 4.0 Hz, J _{1 -3} = 12.0 Hz, 3H), 4.27 (d, J = 8.0 Hz, 6H), 3.72 (t, J = 6.0 Hz, 6H), 3.10 (s, 6H), 1.57 ( quin, J = 8.0 Hz, 6H), 1.29-1.20 (m, 30H), 0.88 (t, J = 6.0 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ): δ 135.5, 116.8, 83.0, 82.6, 74.8, 74.2, 32.0, 30.8, 29.7, 29.5, 26.4, 23.0, 14.3.

<5-2> Synthesis of Compound C9

Compound C9 was synthesized in 73% yield according to the general ozone decomposition reaction procedure of Example 2-2. ¹ H NMR (400 MHz, CDCl ₃ ): δ 3.85-3.83 (m, 6H), 3.74 (t, J = 6.0 Hz, 6H), 3.68 (s, 6H), 3.38 (br s, 3H), 3.25 (t, J = 10.0 Hz, 3H), 3.06 (t, J = 10.0 Hz, 3H), 1.59 (quin, J = 8.0 Hz, 6H), 1.31-1.24 (m, 30H), 0.85 (t, J = 6.0 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ): δ 83.1, 82.9, 74.9, 74.7, 63.4, 32.0, 30.3, 29.7, 29.4, 26.2, 22.8, 14.3.

<5-3> Synthesis of STM-E8a

Compound STM-E8a was synthesized in a yield of 65% according to the general saccharification reaction procedure of Example 2-3. ¹ H NMR (400 MHz, CDCl ₃ ): δ 8.11 (d, J = 8.0 Hz, 6H), 8.01 (d, J = 8.0 Hz, 6H), 7.86 (d, J = 8.0 Hz, 6H), 7.80 ( d, J = 8.0 Hz, 6H), 7.75 (d, J = 8.0 Hz, 12H), 7.65 (d, J = 8.0 Hz, 6H), 7.54 (t, J = 8.0 Hz, 3H), 7.49 (t, J = 8.0 Hz, 3H), 7.46-7.35 (m, 24H), 7.31-7.27 (m, 9H), 7.26-7.21 (m, 18H), 7.15 (t, J = 8.0 Hz, 6H), 6.13 (t , J = 10.0 Hz, 3H), 5.79 (t, J = 8.0 Hz, 3H), 5.77 (t, J = 4.0 Hz, 3H), 5.70 (t, J = 10.0 Hz, 3H), 5.32-5.23 (m , 6H), 4.90 (d, J = 12.0 Hz, 3H), 4.80 (d, J = 4.0 Hz, 3H), 4.74 (dd, J _1-2 = 4.0 Hz, J _1-3 = 12.0 Hz, 3H) , 4.56-4.49 (m, 6H), 4.42 (d, J = 12.0 Hz, 3H), 4.29 (dd, J _1-2 = 4.0 Hz, J _1-3 = 12.0 Hz, 3H), 4.13-4.06 (m , 3H), 3.95 (t, J = 4.0 Hz, 3H), 3.77-3.70 (m, 3H), 3.65-3.63 (m, 6H), 3.47-3.38 (m, 6H), 2.71 (t, J = 12.0 Hz, 3H), 2.47 (t, J = 8.0 Hz, 3H) 1.36-1.32 (m, 6H), 1.25-1.08 (m, 30H), 0.86 (t, J = 6.0 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ): δ 166.2, 165.8, 165.7, 165.5, 165.1, 165.1, 165.0, 133.5, 133.4, 133.3, 133.2, 133.1, 130.0, 129.9, 129.8, 129.7, 129.6, 129.4, 129.3, 128.9, 128.8, 128.7, 128.6, 128.5, 128.4, 128.3, 128.2, 128.1, 101.0, 96.5, 83.0, 82.0, 75.2, 73.6, 73.2, 73.0, 72.3, 71.6, 71.0, 70.9, 70.0, 69.4, 69.2, 63.7, 62.6, 60.4, 53.5, 31.9, 30.4, 29.6, 29.4, 26.3, 22.7, 21.0, 14.2.

<5-4> Synthesis of STM-E8

According to the general synthetic procedure for the deprotection vaporization reaction of Example 2-4, STM-E8 in a yield of 92% Synthesized. ¹ H NMR (400 MHz, CD ₃ OD): δ 5.15 (d, J = 4.0 Hz, 3H), 4.32 (d, J = 8.0 Hz, 3H), 4.05-3.98 (m, 6H), 3.91-3.88 ( m, 6H), 3.83-3.80 (m, 6H), 3.76 (t, J = 6.0Hz, 9H), 3.69-3.61 (m, 12H), 3.52 (t, J = 10.0 Hz, 3H), 3.45 (dd , J _1-2 = 4.0 Hz, J _1-3 = 12.0 Hz, 3H), 3.39-3.35 (m, 3H), 3.28-3.24 (m, 6H), 3.20 (t, J = 10.0 Hz, 3H), 3.09 (t, J = 8.0 Hz, 3H), 1.64-1.62 (m, 6H), 1.39-1.21 (m, 30H), 0.91 (t, J = 8.0 Hz, 9H); ¹³ C NMR (100 MHz, CD ₃ OD): δ 104.4, 103.1, 84.3, 83.8, 81.6, 77.8, 76.8, 75.2, 74.9, 74.8, 74.2, 73.5, 71.5, 70.4, 62.8, 62.3, 33.2, 31.7, 30.9 , 30.6, 27.5, 23.9, 14.7. HRMS (FAB ⁺ ) : calcd. for C ₇₂ H ₁₃₂ O ₃₉ [M+Na] ⁺ 1643.8243, found 1643.8236.

<Production Example 6> Synthesis of STM-E9

<6-1> Synthesis of Compound B6

Compound B6 was synthesized with a yield of 77% according to the general alkylation reaction procedure of 1,3,5-triallylated silitol of Example 2-1. ¹ H NMR (400 MHz, CDCl ₃ ): δ 5.98-5.91 (m, 3H), 5.26 (dd, J _1-2 = 3.0 Hz, J _1-3 = 16.0 Hz, 3H), 5.14 (dd, J _{1 -2} = 4.0 Hz, J _{1 -3} = 12.0 Hz, 3H), 4.27 (d, J = 8.0 Hz, 6H), 3.72 (t, J = 6.0 Hz, 6H), 3.10 (s, 6H), 1.57 ( quin, J = 8.0 Hz, 6H), 1.29-1.20 (m, 36H), 0.88 (t, J = 6.0 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ): δ 135.5, 116.9, 83.0, 82.6, 74.8, 74.3, 32.1, 30.8, 29.8, 29.5, 26.4, 23.0, 14.3.

<6-2> Synthesis of Compound C10

Compound C10 was synthesized in 73% yield according to the general ozone decomposition reaction procedure of Example 2-2. ¹ H NMR (400 MHz, CDCl ₃ ): δ 3.88-3.86 (m, 6H), 3.78 (t, J = 6.0 Hz, 6H), 3.70 (s, 6H), 3.48 (br s, 3H), 3.28 (t, J = 10.0 Hz, 3H), 3.10 (t, J = 10.0 Hz, 3H), 1.63 (quin, J = 8.0 Hz, 6H), 1.31-1.26 (m, 36H), 0.88 (t, J = 6.0 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ): δ 83.0, 82.9, 74.8, 74.7, 63.2, 31.9, 30.2, 29.6, 29.3, 26.1, 22.7, 14.2.

<6-3> Synthesis of STM-E9a

Compound STM-E9a was synthesized in a yield of 65% according to the general saccharification reaction procedure of Example 2-3. ¹ H NMR (400 MHz, CDCl ₃ ): δ 8.08 (d, J = 8.0 Hz, 6H), 7.98 (d, J = 8.0 Hz, 6H), 7.85 (d, J = 8.0 Hz, 6H), 7.77 ( d, J = 8.0 Hz, 6H), 7.73 (d, J = 8.0 Hz, 12H), 7.63 (d, J = 8.0 Hz, 6H), 7.57-7.45 (m, 9H), 7.44-7.37 (m, 21H) ), 7.33-7.29 (m, 9H), 7.27-7.17 (m, 24H), 6.09 (t, J = 12.0 Hz, 3H), 5.75 (t, J = 8.0 Hz, 3H), 5.73 (d, J = 4.0 Hz, 3H), 5.66 (t, J = 10.0 Hz, 3H), 5.30 (t, J = 8.0 Hz, 3H), 5.25 (dd, J _1-2 = 4.0 Hz, J _1-3 = 12.0 Hz, 3H), 4.89 (d, J = 8.0 Hz, 3H), 4.80 (d, J = 8.0 Hz, 3H), 4.75 (dd, J _1-2 = 4.0 Hz, J _1-3 = 12.0 Hz, 3H), 4.50 (t, J = 8.0 Hz, 3H), 4.47-4.43 (m, 3H), 4.37 (dd, J _1-2 = 4.0 Hz, J _1-3 = 8.0 Hz, 3H), 4.24 (dd, J _{1 -2} = 4.0 Hz, J _1-3 = 12.0 Hz, 3H), 4.12-4.07 (m, 3H), 3.93-3.91 (m, 3H), 3.74-3.71 (m, 3H), 3.63-3.58 (m, 6H), 3.43-3.33 (m, 6H), 2.67 (t, J = 10.0 Hz, 3H), 2.45 (t, J = 10.0 Hz, 3H) 1.32-1.08 (m, 36H), 0.86 (t, J = 6.0 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ): δ 166.3, 166.0, 165.8, 165.6, 165.2, 165.1, 133.6, 133.5, 133.4, 133.3, 133.2, 130.1, 130.0, 129.9, 129.8, 129.7, 129.6, 129.5, 129.3, 129.1, 128.9, 128.8, 128.6, 128.5, 128.4, 128.3, 128.2, 101.1, 96.6, 83.1, 82.0, 75.3, 73.7, 73.3, 73.1, 72.3, 71.6, 71.0, 70.1, 69.5, 69.3, 63.7, 62.6, 32.1, 30.6, 29.8, 29.5, 26.5, 22.9, 14.3.

<6-4> Synthesis of STM-E9

According to the general synthetic procedure for the deprotection vaporization reaction of Example 2-4, STM-E9 in a yield of 90% Synthesized. ¹ H NMR (400 MHz, CD ₃ OD): δ 5.15 (d, J = 4.0 Hz, 3H), 4.33 (d, J = 8.0 Hz, 3H), 3.99 (d, J = 0.04 Hz, 6H), 3.86 (d, J = 8.0 Hz, 6H), 3.82-3.75 (m, 15H), 3.70-3.61 (m, 12H), 3.54 (t, J = 10.0 Hz, 3H), 3.47 (dd, J _1-2 = 4.0 Hz, J _1-3 = 12.0 Hz, 3H), 3.39-3.35 (m, 3H), 3.32-3.25 (m, 6H), 3.20 (t, J = 10.0 Hz, 3H), 3.07 (t, J = 8.0 Hz, 3H), 1.64-1.62 (m, 6H), 1.39-1.21 (m, 36H), 0.90 (t, J = 6.0 Hz, 9H); ¹³ C NMR (100 MHz, CD ₃ OD): δ 104.3, 103.1, 84.2, 83.7, 81.5, 77.8, 76.7, 75.1, 74.8, 74.2, 73.4, 71.5, 70.4, 62.8, 62.3, 52.8, 33.2, 31.7, 30.9 , 30.8, 30.6, 27.4, 23.8, 14.7. HRMS (FAB ⁺ ) : calcd. for C ₇₅ H ₁₃₈ O ₃₉ [M+Na] ⁺ 1685.8713, found 1685.8710.

<Production Example 7> Synthesis of STM-E10

<7-1> Synthesis of Compound B7

Compound B7 was synthesized in 75% yield according to the general alkylation reaction procedure of 1,3,5-triallylated silitol of Example 2-1. ¹ H NMR (400 MHz, CDCl ₃ ): δ 6.00-5.90 (m, 3H), 5.25 (d, J = 20.0 Hz, 3H), 5.14 (d, J = 8.0 Hz, 3H), 4.26 (d, J = 4.0 Hz, 6H), 3.72 (t, J = 8.0 Hz, 6H), 3.09 (s, 6H), 1.57 (quin, J = 8.0 Hz, 6H), 1.29-1.20 (m, 40H), 0.88 (t, J = 6.0 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ): δ 135.5, 116.8, 83.0, 82.6, 74.8, 74.2, 32.1, 30.7, 29.8, 29.7, 29.5, 26.4, 22.9, 14.3.

<7-2> Synthesis of Compound C11

Compound C11 was synthesized in 72% yield according to the general ozone decomposition reaction procedure of Example 2-2. ¹ H NMR (400 MHz, CDCl ₃ ): δ 3.88-3.86 (m, 6H), 3.78 (t, J = 6.0 Hz, 6H), 3.70 (s, 6H), 3.48 (br s, 3H), 3.28 (t, J = 10.0 Hz, 3H), 3.10 (t, J = 10.0 Hz, 3H), 1.63 (quin, J = 8.0 Hz, 6H), 1.31-1.26 (m, 40H), 0.88 (t, J = 6.0 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ): δ 83.0, 82.9, 74.8, 74.7, 63.2, 32.0, 30.3, 29.7, 29.6, 29.4, 26.2, 22.8, 14.2.

<7-3> Synthesis of STM-E10a

Compound STM-E10a was synthesized in a yield of 65% according to the general saccharification reaction procedure of Example 2-3. ¹ H NMR (400 MHz, CDCl ₃ ): δ 8.10 (d, J = 8.0 Hz, 6H), 7.99 (d, J = 8.0 Hz, 6H), 7.85 (dd, J _1-2 = 4.0, J _{1- 3} = 12.0 Hz, 6H), 7.78 (d, J = 8.0 Hz, 6H), 7.73 (d, J = 8.0 Hz, 12H), 7.63 (dd, J _1-2 = 4.0, J _1-3 = 12.0 Hz , 3H), 7.55-7.49 (m, 7H), 7.46-7.35 (m, 25H), 7.33-7.27 (m, 12H), 7.25-7.16 (m, 22H), 6.10 (t, J = 12.0 Hz, 3H ), 5.75-5.73 (m, 6H), 5.66 (t, J = 10.0 Hz, 3H), 5.32-5.24 (m, 6H), 4.89 (d, J = 8.0 Hz, 3H), 4.82 (d, J = 8.0 Hz, 3H), 4.75 (dd, J _1-2 = 4.0 Hz, J _1-3 = 12.0 Hz, 3H), 4.50 (t, J = 8.0 Hz, 3H), 4.47-4.40 (m, 3H), 4.37 (d, J = 12.0 Hz, 3H), 4.24 (dd, J _1-2 = 4.0 Hz, J _1-3 = 12.0 Hz, 3H), 4.12-4.07 (m, 3H), 3.93-3.91 (m, 3H), 3.74-3.71 (m, 3H), 3.63-3.58 (m, 6H), 3.43-3.33 (m, 6H), 2.67 (t, J = 10.0 Hz, 3H), 2.45 (t, J = 10.0 Hz , 3H) 1.32-1.08 (m, 40H), 0.88 (t, J = 6.0 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ): δ 166.2, 166.0, 165.8, 165.4, 165.2, 165.1, 133.6, 133.5, 133.4, 133.3, 133.2, 130.1, 130.0, 129.9, 129.8, 129.7, 129.6, 129.5, 129.3, 129.1, 128.9, 128.8, 128.6, 128.5, 128.4, 128.3, 128.2, 101.1, 96.6, 83.1, 82.0, 75.3, 73.6, 73.3, 73.1, 72.3, 71.6, 71.2, 70.1, 69.5, 69.3, 63.7, 62.6, 32.1, 30.6, 29.8, 29.5, 26.5, 22.9, 14.2.

<7-4> Synthesis of STM-E10

According to the general synthetic procedure for the deprotection vaporization reaction of Example 2-4, STM-E10 in a yield of 91% Synthesized. ¹ H NMR (400 MHz, CD ₃ OD): δ 5.15 (d, J = 4.0 Hz, 3H), 4.32 (d, J = 8.0 Hz, 3H), 4.05-3.98 (m, 6H), 3.90-3.88 ( d, J = 12.0 Hz, 6H), 3.85-3.81 (m, 6H), 3.80-3.71 (m, 9H), 3.69-3.60 (m, 12H), 3.53 (t, J = 10.0 Hz, 3H), 3.45 (dd, J _1-2 = 4.0 Hz, J _1-3 = 8.0 Hz, 3H), 3.39-3.35 (m, 3H), 3.32-3.24 (m, 6H), 3.20 (t, J = 8.0 Hz, 3H ), 3.07 (t, J = 8.0 Hz, 3H), 1.64-1.62 (m, 6H), 1.39-1.21 (m, 40H), 0.90 (t, J = 6.0 Hz, 9H); ¹³ C NMR (100 MHz, CD ₃ OD): δ 104.3, 103.1, 84.2, 83.6, 81.5, 77.8, 76.7, 75.1, 74.8, 74.2, 73.4, 71.5, 70.4, 62.8, 62.3, 33.2, 31.7, 30.9, 30.8 , 30.6, 27.4, 23.8, 14.7. HRMS (FAB ⁺ ) : calcd. for C ₇₈ H ₁₄₄ O ₃₉ [M+Na] ⁺ 1727.9182, found 1727.9188.

<Example 3> SHG-Gs Synthesis method

The synthesis scheme of SHG-Gs is shown in FIG. 3. Three kinds of SHG-Gs compounds were synthesized according to the synthesis method of the following <3-1> to <3-5>.

<3-1> 1,3,5- Triallylation -2,4,6- Trialkylated Silitol (1,3,5-triallylated-2,4,6-trialkylated scyllitol ) Synthesis of (Compound B) (Step a in Fig. 3)

NaH (60% dispersion in mineral oil, 6.0 equivalents) was added to 1,3,5-triallylated syllitol (1,3,5-triallylated syllitol) (compound 1b) dissolved in DMF under argon (Ar) atmosphere. I did. Alkyl iodide (4.5 equivalents) was added dropwise after gas evolution ceased, and the reaction mixture was stirred at room temperature for 24 hours and ice was added to quench the reaction. The quenched reaction mixture was diluted with water and then washed with ethyl acetate. The combined ethyl acetate fractions were washed with brine and dried over anhydrous Na ₂ SO ₄ . The solvent was removed under reduced pressure to obtain an oily residue, and purified by column chromatography to give the title compound B as a colored oil.

<3-2> Upzone Dehydroxylation ( UpJohn dihydroxylation ) General synthesis procedure of the reaction (step b in Figure 3)

To a well stirred solution of Compound B in acetone:water:tert-butanol (4:3:1), N-methylmorpholine N-oxide (50 wt% solution, 1.10 mmol per alkene) was added at room temperature. After a few minutes OsO ₄ (0.25 mol% per alkene) was added and stirring was continued at room temperature. After completion of the reaction as indicated by TLC, the solvent was removed under reduced pressure. The resulting brown oily residue was dissolved in DCM, washed with water, brine, and dried over anhydrous Na ₂ SO ₄ . The solvent was removed under reduced pressure to obtain a dark oil, which was purified by column chromatography (SiO ₂ ) to obtain the title compound D.

<3-3> General synthesis procedure of glycosycosylation reaction (step c in FIG. 3)

Under nitrogen (N ₂ ), a mixture of compound D (1.0 equivalent), dimethoxyethane (30.0 equivalent) and AgOTf (7.2 equivalent) was stirred in anhydrous CH ₂ Cl ₂ at -45°C. A solution of perbenzoylated glucosylbromide (7.2 equivalents) in anhydrous CH ₂ Cl ₂ was added dropwise to this suspension. After stirring was continued at -45° C. for several minutes, the temperature of the reaction mixture was increased to 0° C. and stirring was continued for 30 minutes. After completion of the reaction (represented by TLC), pyridine was added to the reaction mixture, diluted with CH ₂ Cl _2, and filtered over Celite. The filtrate was washed successively with 1 M aqueous Na ₂ S ₂ O ₃ solution, 0.1 M HCl aqueous solution, and brine. After drying the organic layer with anhydrous Na ₂ SO ₄ , the solvent was removed using a rotary evaporator. The obtained residue was purified by silica silica gel column chromatography (EtOAc/hexane) to obtain the desired compound D ₁ as a solid.

<3-4> Deprotection vaporization reaction ( deprotection reaction) for general synthesis procedure (step d in FIG. 3)

This was followed by the synthesis method of Chae, PS et al. (Nat Meth 2010, 7, 1003.). De-O-benzoylation was performed under Zemplen's conditions. The O-protected compound was dissolved in anhydrous CH2Cl2 and then MeOH was slowly added until a continuous precipitation appeared. NaOMe, which is a 0.5M methanolic solution, was added to the reaction mixture so that the final concentration was 0.05M. The reaction mixture was stirred at room temperature for 6 hours. After completion of the reaction, the reaction mixture was neutralized using Amberlite IR-120 (H+ form) resin. The resin was removed by filtration, washed with MeOH, and the solvent was removed from the filtrate under vacuum conditions (in vacuo). Diethyl ether (50 mL) was added to the residue dissolved in 2 mL of MeOH:CH ₂ Cl ₂ (1:1) mixture to obtain the target compound D ₂ as a precipitated white solid.

<Production Example 8> Synthesis of SHG-G12

<8-1> Synthesis of Compound B12

Compound B12 was synthesized in 72% yield according to the general alkylation reaction procedure of 1,3,5-triallylated silitol of Example 3-1. ¹ H NMR (400 MHz, CDCl ₃ ): δ 6.00-5.91 (m, 3H), 5.26 (d, J = 16.0 Hz, 3H), 5.14 (d, J = 12.0 Hz, 3H), 4.27 (d, J = 4.0 Hz, 6H), 3.72 (t, J = 6.0 Hz, 6H), 3.10 (s, 6H), 1.57 (quin, J = 8.0 Hz, 6H), 1.29-1.20 (m, 54H), 0.88 (t , J = 8.0 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ): δ 135.5, 116.8, 83.0, 82.6, 74.8, 74.2, 71.1, 32.1, 30.7, 29.9, 29.8, 29.7, 29.6, 26.4, 22.9, 14.3.

<8-2> Synthesis of Compound D15

Compound D15 was synthesized in 82% yield according to the general upzone dehydroxylation reaction procedure of Example 3-2. ¹ H NMR (400 MHz, CDCl ₃ ): δ 4.02 (br s, 1H), 3.89-3.86 (m, 7H), 3.78-3.70 (m, 8H), 3.66 (d, J = 12.0 Hz, 3H), 3.57-3.52 (m, 3H), 3.29-3.23 (m, 3H), 3.10 (quin, J = 8.0 Hz, 3H), 2.42 (br s, 3H), 1.63 (quin, J = 8.0 Hz, 6H), 1.31-1.25 (m, 54H), 0.88 (t, J = 8.0 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ): δ 83.1, 83.0, 82.9, 82.8, 82.7, 82.6, 82.5, 75.1, 74.9, 74.8, 74.7, 74.5, 74.3, 72.1, 72.0, 71.9, 71.0, 66.5, 64.3, 63.4, 55.1, 53.5, 46.1, 32.0, 30.3, 30.2, 29.8, 29.7, 29.6, 29.5, 26.2, 26.1, 25.4, 22.8, 14.2.

<8-3> Synthesis of SHG-G12a

Compound SHG-G12a was synthesized in a yield of 64% according to the general saccharification reaction procedure of Example 3-3. ¹ H NMR (400 MHz, CDCl ₃ ): δ 8.23-7.80 (m, 48H), 7.62-7.23 (m, 72H), 5.86-5.80 (m, 4H), 5.70 (t, J = 10.0 Hz, 3H) , 5.62-5.46 (m, 9H), 5.38-5.28 (m, 2H), 4.90-4.75 (m, 4H), 4.64-4.50 (m, 6H), 4.46-4.44 (m, 4H), 4.38-4.32 ( m, 3H), 4.13-3.98 (m, 9H), 3.72-3.18 (m, 19H), 2.80-2.00 (m, 6H), 1.50-1.40 (m, 3H), 1.38-1.10 (m, 57H), 0.84-0.80 (m, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ): δ 166.1, 166.0, 165.9, 165.8, 165.4, 165.2, 165.1, 164.9, 133.4, 133.2, 130.2, 130.0, 129.9, 129.7, 129.6, 129.5, 129.3, 129.2, 129.1, 128.9, 128.6, 128.5, 128.4, 101.4, 73.3, 73.2, 73.0, 72.8, 72.5, 72.1, 71.9, 71.6, 70.0, 69.7, 63.1, 60.5, 53.6, 32.1, 30.8, 30.2, 30.1, 30.0, 29.9, 29.6, 26.7, 26.6, 22.8, 14.3.

<8-4> Synthesis of SHG-G12

SHG-G12 in a yield of 90% according to the general synthetic procedure for the deprotection vaporization reaction of Example 3-4 Synthesized. ¹ H NMR (400 MHz, CD ₃ OD): δ 4.59 (t, J = 6.0 Hz, 1.5H), 4.49 (d, J = 8.0 Hz, 1.5H), 4.40 (d, J = 8.0 Hz, 1.5H ), 4.33 (d, J = 8.0 Hz, 1.5H), 4.12 (d, J = 8.0 Hz, 2H), 4.06-4.01 (m, 5H), 3.99-3.96 (m, 4H), 3.88-3.80 (m) , 8H), 3.78-3.65 (m, 16H), 3.38-3.28 (m, 9H), 3.28-3.26 (m, 8H), 3.23-3.19 (m, 8H), 3.13-3.09 (m, 3H), 1.69 -1.59 (m, 6H), 1.38-1.20 (m, 54H), 0.90 (t, J = 8.0 Hz, 9H). ¹³ C NMR (100 MHz, CD ₃ OD): δ 105.1, 104.8, 104.6, 84.1, 79.6, 78.2, 78.1, 78.0, 77.9, 75.6, 75.4, 75.3, 75.2, 71.8, 71.7, 71.6, 62.9, 33.2, 31.8 , 31.2, 31.1, 31.0, 30.7, 27.6, 23.9, 14.6. HRMS (FAB ⁺ ) : calcd. for C ₈₇ H ₁₆₂ O ₄₂ [M+Na] ⁺ 1902.0438, found 1902.0431.

<Production Example 9> Synthesis of SHG-G13

<9-1> Synthesis of Compound B13

Compound B13 was synthesized in 72% yield according to the general alkylation reaction procedure of 1,3,5-triallylated silitol of Example 3-1. ¹ H NMR (400 MHz, CDCl ₃ ): δ 6.00-5.90 (m, 3H), 5.26 (dd, J _1-2 = 4.0 Hz, J _1-3 = 20.0 Hz, 3H), 5.14 (dd, J _{1 -2} = 4.0 Hz, J _1-3 = 12.0 Hz, 3H), 4.27 (d, J = 4.0 Hz, 6H), 3.72 (t, J = 6.0 Hz, 6H), 3.10 (s, 6H), 1.56 ( quin, J = 8.0 Hz, 6H), 1.29-1.20 (m, 60H), 0.88 (t, J = 8.0 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ): δ 135.5, 116.8, 82.9, 82.6, 74.8, 74.2, 71.2, 32.1, 30.7, 30.0, 29.9, 29.8, 29.7, 29.6, 26.4, 22.9, 14.3.

<9-2> Synthesis of Compound D16

Compound D16 was synthesized in 82% yield according to the general upzone dehydroxylation reaction procedure of Example 3-2. ¹ H NMR (400 MHz, CDCl ₃ ): δ 4.02 (br s, 1H), 3.86-3.84 (m, 6H), 3.78-3.71 (m, 9H), 3.66 (d, J = 12.0 Hz, 3H), 3.57-3.51 (m, 3H), 3.38-3.23 (m, 3H), 3.14-3.06 (m, 3H), 2.42 (br s, 3H), 1.63 (quin, J = 8.0 Hz, 6H), 1.31-1.25 (m, 60H), 0.88 (t, J = 8.0 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ): δ 83.2, 83.0, 82.8, 82.8, 75.3, 75.1, 75.0, 74.8, 74.5, 72.1, 72.0, 71.9, 71.2, 63.6, 32.1, 30.4, 30.3, 30.2, 30.0, 29.9, 29.8, 29.7, 29.6, 26.4, 26.3, 26.2, 22.9, 14.3.

<9-3> Synthesis of SHG-G13a

Compound SHG-G13a was synthesized in a yield of 63% according to the general saccharification reaction procedure of Example 3-3. ¹ H NMR (400 MHz, CDCl ₃ ): δ 8.25-7.84 (m, 48H), 7.60-7.24 (m, 72H), 5.81-5.77 (m, 4H), 5.67 (t, J = 6.0 Hz, 3H) , 5.64-5.43 (m, 9H), 5.38-5.28 (m, 2H), 4.87-4.70 (m, 4H), 4.60-4.52 (m, 6H), 4.44-4.41 (m, 6H), 4.35-4.27 ( m, 3H), 4.09-3.95 (m, 8H), 3.65-3.15 (m, 20H), 2.66-2.01 (m, 5H), 1.50-1.40 (m, 3H), 1.38-1.10 (m, 63H), 0.84-0.80 (m, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ): δ 166.0, 165.9, 165.7, 165.1, 164.9, 133.4, 133.2, 130.1, 129.9, 129.8, 129.6, 129.4, 129.2, 129.1, 129.0, 128.6, 128.4, 128.3, 101.3, 73.3, 73.0, 72.4, 71.8, 71.5, 70.0, 69.5, 63.0, 41.0, 32.0, 31.9, 30.8, 30.6, 30.2, 30.1, 29.9, 29.8, 29.7, 29.6, 29.5, 29.4, 26.7, 22.9, 22.7, 14.2.

<9-4> Synthesis of SHG-G13

According to the general synthetic procedure for the deprotection vaporization reaction of Example 3-4, SHG-G13 in a yield of 92% Synthesized. ¹ H NMR (400 MHz, sCD ₃ OD): δ 4.59 (t, J = 6.0 Hz, 1.5H), 4.50 (d, J = 8.0 Hz, 1.5H), 4.40 (d, J = 8.0 Hz, 1.5H ), 4.33 (d, J = 8.0 Hz, 1.5H), 4.10 (d, J = 8.0 Hz, 2H), 4.06-4.01 (m, 4H), 3.98-3.96 (m, 5H), 3.88-3.80 (m , 8H), 3.78-3.65 (m, 16H), 3.38-3.28 (m, 8H), 3.23-3.26 (m, 8H), 3.23-3.19 (m, 6H), 3.13-3.09 (m, 3H), 1.69 -1.59 (m, 6H), 1.38-1.20 (m, 60H), 0.90 (t, J = 8.0 Hz, 9H). ¹³ C NMR (100 MHz, CD ₃ OD): δ 105.1, 104.8, 104.6, 84.0, 79.7, 78.1, 78.0, 75.5, 75.2, 71.8, 71.7, 66.3, 63.0, 62.9, 62.6, 33.2, 31.8, 31.1, 31.0 , 30.9, 30.6, 27.6, 23.8, 14.6. HRMS ( FAB ⁺ ) : calcd. for C ₉₀ H ₁₆₈ O ₄₂ [M+Na] ⁺ 1944.0908, found 1944.0921.

<Production Example 10> Synthesis of SHG-G14

<10-1> Synthesis of Compound B14

Compound B14 was synthesized in 71% yield according to the general alkylation reaction procedure of 1,3,5-triallylated silitol of Example 3-1. ¹ H NMR (400 MHz, CDCl ₃ ): δ 6.00-5.92 (m, 3H), 5.26 (d, J = 16.0 Hz, 3H), 5.14 (d, J = 12.0 Hz, 3H), 4.27 (d, J = 8.0 Hz, 6H), 3.72 (t, J = 6.0 Hz, 6H), 3.10 (s, 6H), 1.57 (quin, J = 8.0 Hz, 6H), 1.29-1.20 (m, 66H), 0.88 (t , J = 8.0 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ): δ 135.5, 116.8, 83.0, 82.6, 74.8, 74.2, 32.1, 30.7, 29.9, 29.8, 29.7, 29.6, 26.4, 22.9, 14.3.

<10-2> Synthesis of Compound D17

Compound D17 was synthesized in 81% yield according to the general upzone dehydroxylation reaction procedure of Example 3-2. ¹ H NMR (400 MHz, CDCl ₃ ): δ 3.87-3.85 (m, 6H), 3.77-3.70 (m, 9H), 3.64 (d, J = 12.0 Hz, 3H), 3.55-3.50 (m, 3H) , 3.30-3.24 (m, 3H), 3.15-3.07, 1.63 (quin, J = 8.0 Hz, 6H), 1.31-1.25 (m, 66H), 0.88 (t, J = 8.0 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ): δ 83.1, 83.0, 82.9, 82.8, 82.7, 82.6, 82.5, 75.1, 75.0, 74.9, 74.8, 74.6, 74.3, 72.1, 72.0, 71.9, 66.8, 63.4, 55.3, 46.4, 32.1, 30.3, 30.2, 30.1, 29.9, 29.8, 29.7, 29.5, 26.2, 26.1, 25.1, 22.8, 14.3.

<10-3> Synthesis of SHG-G14a

Compound SHG-G14a was synthesized in a yield of 66% according to the general saccharification reaction procedure of Example 3-3. ¹ H NMR (400 MHz, CDCl ₃ ): δ 8.26-7.84 (m, 48H), 7.59-7.25 (m, 72H), 5.90-5.76 (m, 4H), 5.67 (t, J = 8.0 Hz, 3H) , 5.64-5.43 (m, 9H), 5.34-5.25 (m, 2H), 4.89-4.70 (m, 4H), 4.65-4.49 (m, 6H), 4.47-4.25 (m, 6H), 4.15-3.91 ( m, 11H), 383-3.13 (m, 18H), 2.66-2.02 (m, 5H), 1.49-1.43 (m, 3H), 1.33-1.10 (m, 69H), 0.84-0.80 (m, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ): δ 166.0, 165.9, 165.8, 165.7, 165.3, 165.2, 165.0, 164.8, 133.3, 133.1, 130.1, 129.9, 129.8, 129.7, 129.4, 129.0, 128.6, 128.4, 128.3, 101.3, 72.1, 7.0, 62.8, 60.4, 32.0, 30.7, 30.2, 30.1, 29.9, 29.8, 29.5, 26.7, 22.7, 21.1, 14.2.

<10-4> Synthesis of SHG-G14

SHG-G14 in a yield of 90% according to the general synthetic procedure for the deprotection vaporization reaction of Example 3-4 Synthesized. ¹ H NMR (400 MHz, CD ₃ OD): δ 4.59 (t, J = 10.0 Hz, 1.5H), 4.49 (d, J = 8.0 Hz, 1.5H), 4.41 (d, J = 8.0 Hz, 1.5H ), 4.33 (d, J = 8.0 Hz, 1.5H), 4.12 (d, J = 8.0 Hz, 2H), 4.06-4.01 (m, 5H), 3.99-3.96 (m, 4H), 3.88-3.80 (m) , 8H), 3.78-3.65 (m, 16H), 3.38-3.28 (m, 7H), 3.28-3.26 (m, 10H), 3.24-3.20 (m, 8H), 3.13-3.08 (m, 3H), 1.69 -1.59 (m, 6H), 1.38-1.20 (m, 66H), 0.90 (t, J = 8.0 Hz, 9H); ¹³ C NMR (100 MHz, CD ₃ OD): δ 104.9, 104.7, 104.5, 103.9, 83.9, 79.6, 78.0, 77.9, 77.8, 75.4, 75.3, 75.2, 75.1, 72.2, 71.6, 71.5, 62.9, 62.8, 33.2 , 31.7, 31.0, 31.0, 30.9, 30.6, 27.5, 23.8, 14.7. HRMS (FAB ⁺ ) : calcd. for C ₉₃ H ₁₇₄ O ₄₂ [M+Na] ⁺ 1986.1377, found 1986.1382.

<Experimental Example 1> Characteristics of STMs, STM-Es and SHG-Gs

In order to confirm the properties of STMs, STM-Es and SHG-Gs of Preparation Examples 1 to 10 synthesized according to the synthesis method of Examples 1 to 3, the molecular weight (MW) of STMs, STM-Es and SHG-Gs, The critical micellar concentration (CMC) and the hydrodynamic radii ( R _h ) of the formed micelles were measured.

Specifically, the critical micelle concentration (CMC) was measured using fluorescent dyeing and diphenylhexatriene (DPH), and the hydrodynamic radius ( R _h ) of the micelles formed by each formulation (1.0 wt%) was It was measured through a dynamic light scattering (DLS) experiment. The measured results are shown in Table 1 in comparison with DDM, which is a conventional amphiphilic molecule (detergent).

Detergent MW ^a CMC (mM) CMC (wt%) R _h (nm) ^b ±SD STM-E7 1579.7 ~0.300 ~0.0047 2.5 ±0 .1 STM-E8 1621.8 ~0.150 ~0.0024 3.8 ±0 .2 STM-E9 1663.9 ~0.035 ~0.0006 11.2 ±0 .2 STM-E10 1706.0 ~0.010 ~0.0002 21.1 ±0 .7 STM-11 1615.9 ~0.015 ~0.00024 9.4 ±0 .5 STM-12 1658.0 ~0.010 ~0.00016 12.3 ±0 .1 SHG-G12 1880.2 ~0.015 ~0.0028 2.7 ±0 .2 SHG-G13 1922.3 ~0.012 ~0.0023 2.8 ±0 .1 SHG-G14 1964.4 ~0.008 ~0.0016 2.9 ±0 .1 DDM 510.6 ~0.170 0.0087 3.4 ±0 .1

^a Molecular weight of detergents. ^b Detergent hydrodynamic radius measured at 0.5 wt% detergent concentration by dynamic light scattering (DLS).

The CMC values (0.008 to 0.30 mM) of most STMs, STM-Es and SHG-Gs were significantly smaller compared to the CMC values of DDM (0.17 mM). Therefore, since micelles are easily formed even at low concentrations of STMs, STM-Es and SHG-Gs, the same or superior effect can be exhibited even when using a smaller amount than DDM. In addition, the CMC values of STMs, STM-Es and SHG-Gs decreased as the length of the alkyl chain increased, which is believed to be due to the increase in hydrophobicity as the length of the alkyl chain increased. The size of micelles formed by STMs, STM-Es and SHG-Gs generally tended to increase with increasing alkyl chain length. It is believed that this is because the volume of the head group of the amphiphilic compound is constant, and the volume of the tail group increases as the length of the alkyl chain increases, and the molecular geometry changes from conical to cylindrical. On the other hand, when the alkyl chain length is the same, the larger the size of the hydrophilic head group becomes conical, so the size of the micelles formed becomes smaller (compare SHG-G12 with 6 glucose and STM-12 with 3 maltose). It was confirmed that all SHGs and some STMs formed micelles with a size similar to that of DDM, whereas STMs and STM-Es with a long alkyl chain length formed micelles with a larger size than DDM.

On the other hand, as a result of examining the size distribution of micelles formed by STMs, STM-Es and SHG-Gs through DLS, all SHGs showed only one cluster of micelles, indicating high micelle homogeneity (FIG. 4 ). .

From these results, most of the STMs, STM-Es and SHG-Gs of the present invention have lower CMC values than DDM, so micelles are easily formed even with a small amount, so that the tendency of self-assembly is much greater than that of DDM, STMs, STM-Es And it was confirmed that the size of the micelles formed by SHG-Gs differs according to the relative volumes of the hydrophilic head group and the tail group of the alkyl chain, and that the micelles formed by SHG-Gs have high homogeneity.

< Experimental example 2> STMs , STM- Es And SHG - Gs Dissolved by R. capsulatus superassembly stability evaluation

Engineered Rhodobacter capsulatus ( Rhodobacter capsulatus ) strain expressed in R. capsulatus superassembly was solubilized and purified according to a protocol known in the existing literature (PS Chae, Analyst , 2015, 140, 3157-3163.). A 10 mL aliquot of frozen membrane was thawed and homogenized at room temperature using a glass tissue homogenizer. The homogenate was incubated at 32° C. for 30 minutes with slow stirring. After addition of 1.0 wt% DDM, the homogenate was further incubated at 32° C. for 30 minutes. After ultracentrifugation, the supernatant containing the solubilized LHI-RC (light harvesting complex I and the reaction center) complex was collected, and incubated with Ni ^{2 +} -NTA resin at 4° C. for 1 hour. The resin was added to each of 10 His-SpinTrap columns and washed twice with 500 μL binding buffer (10 mM Tris (pH 7.8), 100 mL NaCl, and 1 x CMC DDM). LHI-RC complex purified by DDM was eluted from the column using a buffer containing 1 M imidazole (2×300 μl). The LHI-RC complex purified by 80 μL of DDM was diluted with 920 μL of the following amphiphilic molecule solution, respectively, to reach a final amphiphilic molecule concentration of CMC + 0.2 wt%; (STMs, STM-Es, SHG-Gs and DDM). The LHI-RC complex produced by each amphiphilic molecule was initially incubated at 25° C. for 10 days, then at 35° C. for 10 days, and then incubated at 45° C. for 10 days. By measuring the absorbance of the sample at 875 nm, protein stability was measured at regular intervals during the 30-day incubation of the protein-amphipathic component sample.

As a result, STMs, STM-Es and SHG-Gs of the present invention were superior to DDM in maintaining the stability of the LHI-RC complex (FIG. 5). When comparing the three groups of amphiphilic compounds, SHG-Gs showed better effects than STMs and STM-Es. It was common for all amphiphilic compounds that the ability to maintain complex stability decreased with increasing temperature.

< Experimental example 3> STMs , STM- Es And SHG - Gs of LeuT Membrane protein Structure stabilization ability evaluation

Thermophilic bacteria Aquifex Aeolicus aeolicus ) derived wild type LeuT (leucine transporter) was purified by the method described previously (G. Deckert et al. Nature 1998, 392, 353-358). LeuT was expressed in E. coli C41 (DE3) transformed with pET16b encoding a C-terminal 8xHis-tagged transporter (expression plasmids provided by Dr E. Gouaux, Vollum Institute, Portland, Oregon, USA) . In summary, after separation of the bacterial membrane and solubilization in 1% (w/v) DDM, the protein was bound to Ni ^{2 +} -NTA resin (Life Technologies, Denmark) and 20 mM Tris-HCl (pH 8.0), 1 mM It was eluted in NaCl, 199 mM KCl, 0.05% (w/v) DDM and 300 mM imidazole. After that, the purified LeuT (about 1.5 mg/ml) was obtained in a buffer equivalent to the above, excluding DDM and imidazole, and the final concentration of STMs, STM-Es, SHG-Gs or DDM was CMC + 0.2% (w/v). It was diluted 10-fold with buffer supplemented with. Protein samples were stored at room temperature for 14 days, centrifuged at regular intervals during incubation, and protein properties were confirmed by measuring [ ³ H]-Leucine binding ability using SPA. SPA was performed with 5 μL of protein samples in buffers at the concentrations mentioned above containing 200 mM NaCl and respective STMs, STM-Es or SHG-Gs (or DDM). The SPA reaction was carried out in the presence of 20 nM [ ³ H]-Leucine and 1.25 mg/ml copper chelate (His-Tag) YSi beads (Perkin Elmer, Denmark). The total [ ³ H]-Leucine binding degree of each sample was measured using a MicroBeta liquid scintillation counter (Perkin Elmer).

As shown in FIG. 6, the initial substrate binding activity of LeuT dissolved in DDM was superior compared to other amphiphilic compounds of the present invention, but it was confirmed that the activity rapidly decreased with time. As a result of comparing long-term substrate-binding activity, all compounds except for SHG-G13 were significantly superior to DDM in maintaining the substrate-binding properties of LeuT during the 14-day incubation period. That is, most of the STMs, STM-Es and SHG-Gs completely retained the transporter substrate binding properties for a long period of time.

< Example 4> STMs , STM- Es And SHG - Gs of MelB _st Membrane protein Structure stabilization ability evaluation

An experiment was conducted to measure the structural stability of MelB _St ( Salmonella typhimurium melibiose permease) protein by STMs, STM-Es and SHG-Gs.

Specifically, the wild type MelB _St (melibiose permease) derived from Salmonella typhimurium having a 10-His tag at the C-terminus using the plasmid pK95ΔAHB/WT MelB _St /CH10 was obtained from the E. coli DW2 strain ( Δ melB and Δ lacZY ). Cell growth and membrane preparation were performed according to the method described in AS Ethayathulla et al . ( Nat. Commun . 2014, 5 , 3009). Protein assays were performed with a Micro BCA kit (Thermo Scientific, Rockford, IL). Membrane sample (final protein concentration of 10 mg/mL) containing MelB _St in solubilization buffer (20 mM sodium phosphate, pH 7.5, 200 mM NaCl, 10% glycerol, 20 mM melibiose) was added to 1.5 wt% DDM, STMs, Mixed with STM-Es or SHG-Gs. The resulting samples were incubated for 90 minutes at 4 different temperatures (0, 45, 55, and 65 °C). Beckman Optima with a TLA-100 rotor at 4°C for 45 minutes to remove insoluble matter. Ultracentrifugation was performed at 355,590 g with a MAX ultracentrifuge. 20 μg of non-ultracentrifuged membrane protein was applied to the untreated membrane or an equal amount of extracts of the above compounds after ultracentrifugation, and the treated samples were loaded into each well in an equal volume. The loaded sample was analyzed by SDS-15% PAGE, and then visualized by immunoblotting with Penta-His-HRP antibody (Qiagen, Germantown, MD).

RSO vesicle manufacturing and Trp→D ² G FRET assay

RSO membrane vesicles were prepared from E. coli DW2 cells containing MelB _St or MelB _Ec through osmotic lysis. In a buffer containing 100 mM KPi (pH 7.5) and 100 mM NaCl, 1.0% DDM and STM-12 were respectively treated with 1.0% DDM and STM-12 in a RSO membrane vesicle at a protein concentration of 1 mg/ml for 60 minutes at 23° C. and a TLA 120.2 rotor Ultracentrifugation was performed at 4° C. for 45 minutes with >300,000 g. FRET (Trp→D ² G) experiments were performed on the supernatant using an Amico-Bowman Series 2 (AB2) Spectrofluorometer. D ² G (2'-( N- Dansyl)aminoalkyl-1-thio-β-D-galactopyranoside, dansylgalactoside) was obtained from Drs. Obtained from Gerard Leblanc and H. Ronald Kaback. D ² G FRET signals were collected at 490 and 465 nm for MelB _St and MelB _Ec , respectively, upon releasing the 290 nm Trp residue. 10 μM D ² G and excess melibiose or equal amounts of water (control) were added to the MelB solution at 1 and 2 minutes, respectively.

As shown in Fig. 7(a), most of STMs and STM-Es excluding SHG-Gs at 0° C. and 45° C. showed similar MelB _st solubilization ability as DDM.

However, at higher temperatures (55° C.), the MelB _st solubilizing potency of DDM fell significantly, while most STMs (especially STM-11 and STM-12) maintained the MelB _st solubilizing ability even at 55° C.

Overall, at low temperatures (0°C to 45°C), DDM showed better protein extraction efficiency than STMs, STM-Es and SHG-Gs, whereas in protein thermal stability experiments measured by increasing the temperature, STMs (especially STM-11 and STM) -12) showed remarkably superior transporter solubilization ability than DDM, confirming that the protein stabilization ability was excellent.

In addition, compared to DDM, STM-12 was confirmed to have excellent protein soluble ability and ability to maintain the function of the membrane protein not only for MelB _St, but also for MelB _Ec , which is a less stable homolog (Fig. 7b). .

< Experimental example 4> STMs , STM- Es And SHG - Gs of β ₂ AR Membrane protein Structure stabilization ability evaluation

₂ 아드레날린성 수용체 (β₂AR), G-단백질 연결 수용체(GPCR) 구조 안정성을 측정하는 실험을 하였다. 즉, DDM으로 정제된 수용체는 CHS (cholesteryl hemisuccinate) 없이 각각의 STMs, STM-Es 및 SHG-Gs 만을 함유하는 버퍼 용액 또는 CHS와 DDM을 함유하는 버퍼 용액으로 희석시켰다. 최종 화합물 농도는 CMC+0.2 wt%이었으며, 수용체의 리간드 결합 특성은 [³H]-디하이드로알프레놀올 ([³H]-DHA)의 결합에 의해 측정하였다.STMs, STM-Es and SHG-Gs By human

₂ Experiments were conducted to measure the structural stability of adrenergic receptor (β ₂ AR) and G-protein coupled receptor (GPCR). That is, the receptors purified with DDM are each of STMs, STM-Es and SHG-Gs without CHS (cholesteryl hemisuccinate). It was diluted with a buffer solution containing mannose or a buffer solution containing CHS and DDM. Final compound concentrations were CMC + 0.2 wt%, the ligand binding characteristics of the receptor is ^[3 H] - were measured by a combination of dihydro alpeure nolol ^([3 H] -DHA).

Specifically, the radioligand binding test used the following method: β ₂ AR was purified using 0.1% DDM (DM Rosenbaum et al., Science , 2007, 318, 1266-1273.), about 10 mg/ml (About 200 μM) was finally concentrated. Contains 10 nM [ ³ H]-Dihydroalprenolol (DHA) supplemented with 0.5 mg/ml BSA in 0.2% amphiphilic compounds (DDM, STMs, STM-Es and SHG-Gs) using DDM purified β ₂ AR A master binding mixture was prepared. The receptor purified by DDM, STMs, STM-Es and Gs-SHG was incubated with ^[3 H] -DHA of 10 nM for 30 minutes at room temperature. The mixture was loaded on a G-50 column, and the flow-through was collected with 1 ml binding buffer (20 mM HEPES pH 7.5, 100 mM NaCl supplemented with 0.5 mg/ml BSA and 20xCMC respective amphiphilic compounds), and 15 ml Filled with scintillation fluid. Receptor-bound [ ³ H]-DHA was measured with a scintillation counter (Beckman). Non-specific binding of the ^[3 H] -DHA was measured by the addition of alpeure nolol (alprenolol, Sigma) of 2μM in the same binding reaction. The degree of binding of [ ³ H]-DHA was shown in a column graph, and each experiment was performed three times.

As shown in Figure 8, in maintaining the ligand binding properties of the receptor, STM-E7 showed a similar or superior effect to DDM in terms of initial activity and long-term activity, whereas most other amphiphilic compounds were slightly lower than DDM. Showed activity.

Claims (17)

A compound represented by the following Formula 1 or Formula 2:
[Formula 1]

In Formula 1,
Each of R ¹ to R ³ is independently a substituted or unsubstituted C ₃ -C ₃₀ alkyl group;
Wherein L ¹ to L ³ are each independently a direct bond or -O(oxygen)-Y-, wherein Y is a substituted or unsubstituted C ₁ -C ₁₀ alkylene group; And
X ¹ to X ³ are saccharides linked to oxygen.
[Formula 2]

In Chemical Formula 2,
Each of R ¹ to R ³ is independently a substituted or unsubstituted C ₃ -C ₃₀ alkyl group;
L ⁴ to L ⁶ are substituted or unsubstituted C ₁ -C ₁₀ alkylene groups; And
X ⁴ to X ⁹ are saccharides linked to oxygen.
The compound according to claim 1, wherein the saccharide is a monosaccharide or a disaccharide.
The compound of claim 1, wherein the saccharide is glucose or maltose.
The method of claim 1, wherein R ¹ to R ³ are each independently a substituted or unsubstituted C ₃ -C ₂₀ alkyl group; X ¹ to X ⁹ are glucose or maltose linked to oxygen; In Formula 1, L ¹ to L ³ are a direct bond or -O-CH ₂ CH ₂ -; And L ⁴ to L ⁶ in Formula 2 is -CH ₂ -.
The compound of claim 1, wherein the compound is one of the following Formulas 3 to 12:
[Formula 3]

[Formula 4]

[Formula 5]

[Formula 6]

[Formula 7]

[Formula 8]

[Formula 9]

[Formula 10]

[Formula 11]

[Formula 12]

The compound of claim 1, wherein the compound has a critical micelle concentration (CMC) of 0.0001 to 1 mM in an aqueous solution.
A composition for dissolving a membrane protein comprising the compound according to claim 1.
A composition for extraction of membrane proteins comprising the compound according to claim 1.
A composition for stabilizing a membrane protein comprising the compound according to claim 1.
A composition for crystallization of a membrane protein comprising the compound according to claim 1.
A composition for analysis of membrane proteins comprising the compound according to claim 1.
13. The composition according to any one of claims 7 to 12, wherein the composition is a formulation of micelles, liposomes, emulsions or nanoparticles.
The method of any one of claims 7 to 12, wherein the membrane protein is LHI-RC, LeuT (Leucine transporter), β ₂ AR (human β ₂ adrenergic receptor), MelB _st (melibiose permease), or two or more thereof. A composition that is a combination.
1) performing an alkylation reaction by adding alkyl iodide to silitol-1, 3, 5-orthoacetate;
2) treating the product of step 1) with an acid to remove the orthoacetate protecting group to produce a triol;
3) introducing a saccharide having a protecting group attached thereto by performing a glycosylation reaction to the product of step 2); And
4) performing a deprotection reaction on the product of step 3); including, a method for preparing a compound represented by the following formula (1):
[Formula 1]

In Formula 1,
Each of R ¹ to R ³ is independently a substituted or unsubstituted C ₃ -C ₃₀ alkyl group;
L ¹ to L ³ are direct bonds; And
X ¹ to X ³ are saccharides linked to oxygen.
1) performing an alkylation reaction by adding an alkyl iodide to 1,3,5-triallylated scyllitol;
2) performing an ozonolysis reaction on the product of step 1);
3) introducing a saccharide to which a protecting group is attached by performing a glycosylation reaction to the product of step 2; And
4) performing a deprotection reaction on the product of step 3); including, a method for preparing a compound represented by the following formula (1):
[Formula 1]

In Formula 1,
Each of R ¹ to R ³ is independently a substituted or unsubstituted C ₃ -C ₃₀ alkyl group;
L ¹ to L ³ are -O(oxygen)-Y-, wherein Y is a substituted or unsubstituted C ₁ -C ₁₀ alkylene group; And
X ¹ to X ³ are saccharides linked to oxygen.
1) Step of performing an alkylation reaction by adding an alkyl iodide to 1,3,5-triallylated scyllitol
2) performing dihydroxylation on the product of step 1);
3) introducing a saccharide to which a protecting group is attached by performing a glycosylation reaction to the product of step 2; And
4) performing a deprotection reaction on the product of step 3); including, a method of preparing a compound represented by the following formula (2):
[Formula 2]

In Chemical Formula 2,
Each of R ¹ to R ³ is independently a substituted or unsubstituted C ₃ -C ₃₀ alkyl group;
L ⁴ to L ⁶ are substituted or unsubstituted C ₁ -C ₁₀ alkylene groups; And
X ⁴ to X ⁹ are saccharides linked to oxygen.
A method for extracting a membrane protein, comprising treating the membrane protein with a compound represented by the following Formula 1 or Formula 2 in an aqueous solution:
[Formula 1]

In Formula 1,
Each of R ¹ to R ³ is independently a substituted or unsubstituted C ₃ -C ₃₀ alkyl group;
Wherein L ¹ to L ³ are each independently a direct bond or -O(oxygen)-Y-, wherein Y is a substituted or unsubstituted C ₁ -C ₁₀ alkylene group; And
X ¹ to X ³ are saccharides linked to oxygen.
[Formula 2]

In Chemical Formula 2,
The R ¹ to R ³ are each independently a substituted or unsubstituted C ₃ -C ₃₀ alkyl group,
; And
X ⁴ to X ⁹ are saccharides linked to oxygen.

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