JPS62228296A - Production of peptide or derivative thereof - Google Patents

Abstract

PURPOSE:To remarkably improve the reaction rate within a high pH region, by reacting an alpha-amino acid with an amino acid (derivative) in the presence of an aminoacyl-tRNA synthetase immobilized on a water-insoluble carrier. CONSTITUTION:An alpha-amino acid is reacted with an amino acid or derivative thereof in the presence of an aminoacyl-tRNA synthetase immobilized on a water-insoluble carrier. Examples of the water-insoluble carrier include derivatives of polysaccharides, e.g. cellulose, dextran, etc., polycellulose acetate, polyvinyl alcohol derivatives, etc. In order to immobilize the aminoacyl-tRNA synthetase on the water-soluble carrier, linking by a method using a well-known covalent bonding method, ionic bonding method, etc., or adsorption by a physical adsorption method, entrapping method, etc., may be adopted.

Description

DETAILED DESCRIPTION OF THE INVENTION (Industrial Field of Application) The present invention relates to a method for producing a peptide or a peptide derivative.

(Prior Art) In recent years, it has become increasingly known that peptides have various physiological activities, and their importance as pharmaceuticals for treatment and diagnosis as well as as taste substances is increasing. Along with this, development of peptide synthesis methods is also active. The main methods of peptide synthesis known to date are 1. For example, as summarized in Pharmacia, Review, No. 3, pp. 27-47 (1980), there are two methods: chemical synthesis and enzymatic methods. It can be roughly divided into Typical chemical synthesis methods include the azide method, mixed acid anhydride method, active ester method, and carbodiimide method in which amino acids are condensed sequentially and fragments. The synthetic method also has various problems such as racemization and side reactions are likely to occur, two reaction times are long, and the terminal amino group must be protected with a protecting group before the reaction. For the fragment condensation method.

It has a serious drawback that racemization is particularly likely to occur.

On the other hand, as a way to avoid racemization as much as possible.

An enzymatic method using protease has been proposed.

Even in this method, the reaction time was long, and the terminal amino group had to be protected with a protecting group, so that the complexity of the operation could not be improved.

Furthermore, in the enzymatic method using this protease.

Since the enzyme used originally has peptidolytic activity, the resulting peptide is degraded while being synthesized.

A serious drawback was that the desired peptide was often not obtained. In particular, it has been pointed out that when oligopeptide synthesis is applied, a serious drawback is that unintended peptides lacking some amino acids can be obtained [Journal of Biological Chemistry, 256
Volume, 1301 pages (1981)]. In addition to the protease method, methods for synthesizing peptides using enzymatic methods include methods using special enzymes that control only the synthesis of a single peptide having a specific amino acid sequence. Examples of this type of enzyme include glutathione synthase (Japanese Unexamined Patent Publication No. 122793/1983), which synthesizes a tripeptide with the sequence glutamic acid/cystine/glycine, and gramicidin S synthase, which synthesizes gramicidin S, a decapeptide. (Gendai Kagaku, December 1974 issue, p. 12) have been reported. However, these enzymes are special enzymes.

The peptides that can be synthesized by this enzyme are limited to only one type, and it is not possible to synthesize any desired peptide. For this reason.

At present, this method cannot be used as a general peptide synthesis method.

In view of the usefulness of peptides, the present inventors sought to solve the above-mentioned drawbacks, particularly the need for protective groups that cause racemization, side reactions, and complexity of two reactions, and at the same time impair economic efficiency. As a result of extensive research with the aim of providing a new and versatile peptide synthesis method, we have discovered that an enzyme that has the ability to bind amino acids to tRNA, a type of nucleic acid, has not previously had the ability to form peptide bonds at all. They surprisingly discovered that a previously unknown aminoacyl-t RNA synthetase has the ability to synthesize peptides, and discovered that all of the above objectives could be achieved by using this enzyme as a condensing agent, and filed a patent application earlier. (Unexamined Japanese Patent Publication No. 58-146539 and Unexamined Japanese Patent Publication No. 59-106298)
(see publication).

(Problems to be solved by the invention) This Japanese Patent Application Laid-Open No. 58-146539 and Japanese Patent Application Laid-Open No. 59-1
The method described in JP 06298 is because the optimum pH of aminoacyl-t RNA synthetase used as a condensing agent is 8 to 9.

When the pH of the reaction is 9 or higher, aminoacyl-LRNA
A decrease in the catalytic activity of the synthetase occurred, and conversely, when the pH was lowered to below 8, a similar decrease in the catalytic activity occurred, and the amount of peptide or peptide derivative produced per unit time tended to decrease.

(Means for Solving the Problems) Therefore, the inventors of the present invention conducted further intensive research to improve the above points, and surprisingly found that aminoacyl-
When tRNA synthetase is immobilized on a water-insoluble carrier,
The present invention has been completed based on the discovery that the catalytic ability of aminoacyl-t RNA synthetase is enhanced, and the reaction rate is significantly increased, especially in a high pH region.

That is, the present invention provides a method for producing a peptide or a peptide derivative by reacting an α-amino acid with an amino acid or an amino acid derivative derived from the amino acid in the presence of an aminoacyl-tRNA synthetase.
This is a method for producing a peptide or a peptide derivative, characterized in that an aminoacyl-tRNA synthetase immobilized on a water-insoluble carrier is used as the tRNA synthetase.

The aminoacyl-t RNA synthetase used in the present invention belongs to enzyme classification 6.1.1 and has the primary formula amino acid+A
TP + tRNA - An enzyme that catalyzes the reaction of aminoacyl-t RNA + AMP + birophosphate. For example, rabbit.

Those obtained from animal tissues such as horses, cows, rats, chickens, and snakes; those obtained from plant tissues such as rice, potatoes, and tomatoes; those obtained from microorganisms such as molds, yeasts, mushrooms, bacteria, actinomycetes, and algae. Things can be given. Among these, enzymes obtained from microorganisms are preferred because they are easy to obtain, and Bacillus stearothermophilus, Thermus thermophilus, Thermus flavus, and Stridium thermoaceticum are preferred because of the stability of the enzyme. Aminoacyl-tRNA synthetases obtained from thermostable bacteria such as , Thermus aquaticus and the like are most suitable.

Among these various aminoacyl-t RNA synthetases, those with specificity for various α-amino acids are used.
For example, tyrosyl-tRNA synthetase is specific for tyrosine.

In addition, leucine-specific substances include leucyl-
Examples of valine-specific tRNA synthetase include valyl-tRNA synthetase, isophthalate tRNA synthetase, phenylalanyl-tRNA synthetase, alanyl-tRNA synthetase, glutaminyl-tRNA synthetase, asparaginyl-1RNA synthetase, and methionyl-tRNA synthetase. -tR
NA synthetase, histidyl-t RNA synthetase.

Lysyl-t RNA synthetase, threonyl-t1?
NA synthetase, seryl-t RNA synthetase,
asparatyl-t RNA synthetase, glutamyl-t
tRNA synthetase, cysteinyl-tRNA synthetase, prolyl-tRNA synthetase, glycyl-tRNA synthetase.

Specific examples include arginyl-tRNA synthetase and tryptophanyl-tRNA synthetase.

These various aminoacyl-t RNA synthetases are prepared by disrupting the above-mentioned tissues or cells with a homogenizer or Dynomyl, etc.1, for example, Biochemistry Magazine, 13
Vol., p. 2307 (1974), chromatography such as DEAE-cellulose column chromatography, hydroxyapatite column chromatography, and fractional precipitation with ammonium sulfate.

It can be obtained by purification using a conventional enzyme purification method.

In the present invention, these aminoacyl-t RNA synthetases need to be immobilized on a water-insoluble carrier.

The water-insoluble carrier as used in the present invention essentially refers to a compound that is insoluble in water, and such compounds include, for example, derivatives of multi-tJMs such as cellulose, dextran, agarose, and starch, and polycellulose acetate. , polyvinyl alcohol derivatives, polystyrene, polypropylene,
Polyethylene, polyvinyl chloride, poly(methyl methacrylic acid) ester, polybutene, polyvinylidene chloride, polyacrylonitrile, polymethacrylic acid.

Polyacrylic acid, polyaminostyrene, polybutadiene, polyisoprene, polymaleic acid monoester, crosslinked polyacrylamide, polymethacrylamide, polyvinylamine, poly(dialkylamine ethyl methacrylate), poly(dialkylaminomethylstyrene), poly(vinylpyridine) ), poly(vinylpyrrolidone), polyacrylic anhydride, polymethacrylic anhydride, polymaleic anhydride, polymethacrylic nitrile, poly(trifluoroethylene), poly(tetrafluoroethylene), poly(divinylbenzene), poly
(α-methylstyrene), poly-(N-vinylamine)
, poly(tetramethylene glycol divinyl ether), polyvinyl sulfone, polyvinyl sulfoxide,
Polymers made of monomers containing unsaturated carbon such as polyacrolein and polymethyl vinyl ketone; polyethers such as polyphenylene oxide, polymethylene oxide, polyethylene oxide, and polytetramethylene oxide;
Polypeptides such as polyalanine and polyphenylalanine, nylon-3, nylon-4, nylon-5, nylon-6, nylon-7, nylon-1), nylon-12, nylon-6, 6, nylon-6, 10. Polyamides such as poly(...-phenylene-isophthalamide) and poly(P-phenylene terephthalamide), terephthalic acid, and isophthalic acid.

Polycarboxylic acids such as adipic acid, maleic acid, fumaric acid, and trimellitic acid, as well as ethylene glycol, propylene glycol, and butylene glycol.

Polyesters derived from polyols such as pentaerythritol and bisphenol-A.

Polyesters derived from glycolic acid, lactic acid, hydroxyhybaric acid, etc., silicone rubbers such as dimethylpolysiloxane, methylphenylpolysiloxane, methylvinylpolysiloxane, cyanoalkylmethylpolysiloxane, fluoroalkylmethylpolysiloxane, toluene diisoanate, xylene diisocyanate, phenylene diisocyanate, ethylene diisocyanate,
Polyurethanes derived from polyisocyanates such as diphenylmethane diisocyanate and toluene diisocyanate and polyols such as polyethylene glycol, polypropylene glycol, and polyesters having OH at both ends Phenol-formaldehyde resin, xylene-formaldehyde resin, urea -Formaldehyde resin.

Formaldehyde resins such as melamine-formaldehyde resins, polymers containing four-membered rings such as polyimide, polybenzimidazole, and polythiazole, synthetic polymers such as polycarbonate and polysulfone, glass, asbestos, clay 3 mica, hydroxyapatite, activated carbon, and silica gel.

Examples include inorganic derivatives such as alumina and synthetic inorganic polymers such as polyphosphazene.

In the present invention, to obtain an immobilized aminoacyl-tRNA synthetase, the aminoacyl-tRNA synthetase described above may be bound to or adsorbed onto a water-insoluble carrier. In order to bind aminoacyl-t RNA synthetase to a water-insoluble carrier, for example,
The conventionally known covalent bonding method as described in "Immobilized Enzyme" Kodansha (1975) and 1. Ionic bonding methods can be employed. Further, for adsorption, a physical adsorption method or an entrapment method can be similarly adopted.

As a covalent bonding method, for example, a carrier containing a carboxyl group may be used with azide, chloride, or carbodiimide.

A method in which the derivative is made into a derivative such as an isocyanate and then combined with aminoacyl-tRNA synthetase.

A method of binding an acynoacyl-tRNA synthetase to a water-insoluble carrier having an active leaving group such as a halogen; a method of binding an aminoacyl-tRNA synthetase after treating a water-insoluble carrier having a hydroxyl group with cyanogen halide; A water-insoluble carrier having an aromatic amino group is used as a diazonium salt, and this and aminoacyl-tRN
A method of diazo coupling with A synthetase, using at least a bifunctional reagent such as toluene diisocyanate, epichlorohydrin, glutaraldehyde, 2-amino 4,6-dichloro-8-triazine, etc., to a water-insoluble carrier and an aminoacyl- Examples include a method of binding tRNA synthetase.

Examples of the ionic bonding method include carboxymethylcellulose, diethylaminoethyl Sephadex (Pharmacia), DOWEX-50 (Dow Chemical)
Examples include a method of ionically binding aminoacyl-tRNA synthetase to an ion exchanger such as.

Examples of physical adsorption methods include methods in which aminoacyl-tRNA synthetase is adsorbed onto activated carbon, alumina, silica gel, and the like.

Further entrapment methods include, for example, methods in which aminoacyl-tRNA synthetase is encased in a lattice of a polymer gel such as cross-linked polyacrylamide gel or polyvinyl alcohol, or in a film of nylon, polyester, polystyrene, collodion, or the like.

These immobilization methods can also be carried out by a known method in which an aqueous solution of aminoacyl-tRNA synthetase and a water-insoluble carrier or its precursor are mixed before the initiation of polymerization.

The immobilized aminoacyl-tRNA synthetase used in the present invention can be produced by two different methods as described above, but the binding strength between the aminoacyl-tRNA synthetase and the water-insoluble carrier is large, and therefore the sustainability of the activity is low. It is particularly desirable to produce by a covalent bonding method because it is excellent in that, for example, the synthesis reaction of peptides or peptide derivatives can be carried out stably over a long period of time.

Examples of α-amino acids used in the present invention include tyrosine, alanine, leucine, isoleucine, phenylalanine, methionine, and lysine.

Serine 9 Valine, Asparagine, Aspartic acid.

Glycine, glutamine, glutamic acid, cysteine, threonine, tryptophan, histidine.

Examples include amino acids such as proline and arginine.

Either L-form or D-form may be used. Examples of amino acids or amino acid derivatives to be reacted with α-amino acids include glycine, alanine, and leucine.

Isoleucine, phenylalanine, glutamine.

cysteine, tyrosine, arginine, valine, lysine,
histidine, aspartate, methionine.

α-amino acids such as tryptophan and threonine.

β-amino acids such as β-alanine and β-aminoisoacetic acid, and nitrogen-containing γ-amino acids such as creatine.

Examples include various amino acids such as T-amino acids such as piperidic acid, ε-amino acids such as ε-aminocaproic acid, and esters, thioesters, amides, and hydroxamides of these various amino acids.

The compounds are not limited to the above-mentioned exemplified compounds as long as they are amino acid derivatives in which the amino group is in a free form.

Examples of the ester include methyl and ethyl.

From simple hydrocarbon esters such as propyl, cyclohexyl, phenyl, and benzyl, the 3'-
Two kinds of esters can be used, including those obtained by esterifying the above amino acids with OH.

Furthermore, as the amide, in addition to a free amide, for example, an oligopeptide or polypeptide in which different or the same type of amino acid is amide-bonded can also be used. These oligopeptides and polypeptides are further processed into esters, thioesters, and hydroxamides.

It is also possible to use an etherified one.

According to the present invention, an α-amino acid, an amino acid or an amino acid derivative, an aminoacyl-tR immobilized on a water-insoluble carrier
NA synthetase (hereinafter referred to as immobilized aminoacyl-tRNA)
It is called synthetase. ) and nucleoside triphosphates and reacting them in a liquid phase medium to produce a peptide or peptide derivative. This nucleoside triphosphate is a compound that serves as an energy source for proceeding with two reactions, and a specific example of such a compound is:

Adenosine triphosphate, 2'-deoxyadenosine triphosphate, 2',3'-dideoxyadenosine triphosphate, 3
Examples include '-deoxyadenosine triphosphate, β or T-thio analogs of adenosine triphosphate, and adenosine triphosphate with a substituent on the adenine ring. At this time,
These raw materials may be added in any order;
alpha-amino acid.

The immobilized aminoacyl-tRNA synthetase may be added to an aqueous medium containing the amino acid or amino acid derivative and the nucleoside triphosphate, or the α-amino acid, amino acid or amino MHM4 may be added to the solution containing the immobilized aminoacyl-tRNA synthetase. and nucleoside triphosphates may also be added. Further, the concentration of these raw materials is not particularly limited, but the concentration of α-amino acid is suitably 0.1 mM or more, preferably 1 mM or more. Then, 1 to 15 equivalents of nucleoside triphosphate are added to the α-amino acid, and the immobilized aminoacyl-LRNA synthetase is added to the α-amino acid at 1 to 15 equivalents.
/1 to 1/10 g equivalent, preferably 1/10'' to 1/10 g equivalent.

Amino acid or amino acid derivative 2 usually 10mM to I
It is preferable to add it within the range of OM.

As the medium used for the reaction at this time, a solvent containing water as a main component is selected since the two-prong method is a reaction using an enzyme as a catalyst. Furthermore, a water-soluble organic solvent may be added to the extent that enzyme activity can be maintained. Examples of water-soluble organic solvents include methanol, ethanol, acetonitrile, dioxane, tetrahydrofuran, N,N-dimethylacetamide, N-methylpyrrolidone, and dimethylsulfoxide. Addition of such an organic solvent is particularly effective when some of the raw materials are poorly soluble in water. At this time, divalent cations such as magnesium and manganese, and mercaptoethanol are added to the reaction system for the main purpose of making the reaction proceed smoothly or preventing enzyme deactivation.

Sulfhydryl agents such as dithiothreitol and pyrophosphatase may be added alone or in combination. The preferred concentration of each additive is 0.01mM to 0.01mM of divalent cation.
500mM, sulfhydryl agent 0.001mM to 100
mM, pyrophosphatase 0, OOl nift/m
1 to 1 OO units/m 1 .

The optimal concentration is 0.1mM of each divalent cation.
~10mM, sulfhydryls 0.01mM to 1mM
, pyrophosphatase 1) nift/m (1-10
12・-) ) / m 1. Further, in order to maintain the activity of the enzyme, it is preferable to add a buffer to the solvent.

The concentration of the buffer solution is preferably 100 mM or less. This buffer solution should dissolve α-amino acids, amino acids or amino acid derivatives, immobilized aminoacyl-tRNA synthetase, and nucleoside triphosphates, maintain enzyme activity, obtain the desired pII, and avoid side reactions. Any material may be used as long as it does not cause the problem. Examples of such are Hepes laxative solution, triethanolamine buffer, malate buffer, phosphate buffer.

Bicine slowing solution, Epps slowing solution, boric acid buffer.

Examples include sodium carbonate buffer, sodium hydrogen carbonate buffer, sodium borate buffer, and sodium hydroxide buffer.

In addition, the pH of the reaction solution when producing the peptide or peptide derivative is
When RNA synthetase is not immobilized on a water-insoluble carrier, the optimum pH for the reaction is usually around 8 or 9, but for immobilized aminoacyl-tRNA synthetase, the catalytic activity is significantly increased at alkaline pH of 9 or above. It can be maintained at 9 or above, preferably between 12 and 12, with the buffer mentioned above. The reaction temperature is not particularly limited as long as the catalytic activity of the immobilized aminoacyl-tRNA synthetase can be maintained, but it is usually preferably 0 to 70°C, and most preferably 20 to 60°C.

Under the above conditions, the peptidation reaction is completed in a few seconds to several days, and the desired peptide or peptide derivative can be obtained.

(Example) The present invention will be specifically explained below using examples.

The aminoacyl-tRNA synthetase activity in Examples was determined as follows.

Namely, in the presence of 167mM Hepes buffer, 17mM disodium adenosine triphosphate, 17mM magnesium chloride, 5mM dithiothreitol, 1M hydroxylamine and 5mM α-amino acids.
The ability of lnmde to form α-amino acid hydroxamate when reacted at ℃ for 10 minutes was defined as 1 unit.

Reference Example 1 6 kg of bacterial cells of Bacillus stearothermophilus UK788 (Feikoken Kyori No. 5141) were added to 100
After suspending the cells in mM Tris/HCl buffer (pH 7.5) and disrupting the cells using Dynomill, insoluble materials were removed by centrifugation to obtain a crude extract containing tyrosine-specific tyrosyl-1 RNA synthetase. . A column filled with Madrex Gel Blue A (manufactured by Amicon) equilibrated in advance with 50 mM Lys buffer (pH 7.5) containing 5 mM mercaptoethanol, 2 mM sodium ethylenediaminetetraacetate, and 0.1 mM phosphophenylsulfonyl fluoride. , through the above crude extract.

A solution of potassium chloride added to the above buffer solution at a linear velocity of 60
When eluted with cm.tl, tyrosyl-tRNA synthetase was eluted. Collect this classification.

As a result of concentration and desalting, 1,400,000 units of tyrosine-specific tyrosyl-tRNA RNA synthetase crude enzyme solution was obtained with a yield of about 70%. The above 12 crops were all carried out at 4°C.

Reference example 2 Satucharomyces cerevisiae αS288C1000k
After cell disruption of g with Dynomill, the resulting crude extract was subjected to ammonium sulfate fractionation, DEAE--& Reulose chromatography, cellulose phosphate (Whatman) chromatography, DEAE-Sephacel (Pharmacia) chromatography, and Ultrogel. 3.2 g (5,000
,000 units).

Example 1 Specific activity obtained in Reference Example 1: 6,6001. -F) / me
50mM of tyrosyl-t RNA synthetase
To 30 ml of potassium phosphate buffer was added 2 g of Pharmcyan-activated Sepharose 4B (manufactured by Pharmacia), and the mixture was stirred at room temperature for 4 hours at 4° C. to obtain a specific activity of 1).
,0001: Futo/? An immobilized tyrosyl-t RNA synthetase (hereinafter referred to as immobilized enzyme) exhibiting W heavy N (g) was obtained.

This immobilized enzyme or unimmobilized free enzyme (
Hereinafter referred to as free enzyme. ) 40,000 units contains 18 mg of L-tyrosine and 81 mg of magnesium chloride hexahydrate.
mg, adenosine triphosphate disodium 220mg, L
- Add 421 mg of algine hydrochloride, make the total volume 20 ml with the buffer solution (50 mM) listed in Table 1, and store at 45°C for 2 hours.
The mixture was stirred for several days.

The resulting reaction solution was analyzed by high performance liquid chromatography using a Tsubapak CLS column (manufactured by Waters), and the amount of tyrosylargine produced was as shown in Table 1, indicating that tyrosyl-t RNA synthetase By immobilizing.

A significant increase in yield was observed in the high pH region.

Further, the obtained reaction solution was treated with an aqueous ammonium hydrogen carbonate solution (pH 7,9) using a carboxymethyl cellulose column.
The product was purified by elution with a gradient from 5mM to 100mM. NMR showed that the obtained product was tyrosylargine.

Confirmed by UV and IR absorption spectra.

Table 1 Example 2 Specific activity obtained in Reference Example 2 14.0001: t)/n+j2
Leucyl-tRNA synthetase in 50mM potassium phosphate solution 10+yj! and acrylamide monomer 7
．． 5g, N,N'-methylenebisacrylamide 40m
g and water were mixed to make the total amount 30+yj2. Add water to 250 mg of β-dimethylaminopropionitrile and 50 mg of ammonium persulfate, add 50 m/ml of the solution, mix well, and react at 0°C for 10 minutes to obtain a specific activity of 18001 nif)/m(l). immobilized leucyl-t RNA synthetase (hereinafter referred to as immobilized enzyme).

) was obtained.

Next, L-leucine 13 mg, adenosine triphosphate disodium 1) 0 mg, magnesium chloride hexahydrate 40 mg
, 164 mg of L-phenylalanine and 10,000 units of the above immobilized enzyme or free enzyme were added.
The total volume was made up to 10 ml with 1 (500 mM) of the buffer listed in the table, and the reaction was carried out at 45° C. for 2 days.

The resulting reaction solution was subjected to high performance liquid chromatography in the same manner as in Example 1 to measure the amount of leucylphenylalanine produced.

The results are shown in Table 2.

Table 2 Example 3 Using coconut shell charcoal of 32-60 mesh, Biotechnology and Bioengineering Journal Vol. 24, 1
After reduction with lithium aluminum hydride according to the method described on page 653 (1982), granular coconut shell charcoal containing cyanuric chloride as an active group on the surface was obtained by reacting with cyanuric chloride.

18000 x Nift/for 50mg of this coconut husk charcoal
Add 500 μl of a tyrosyl-tRNA synthetase solution with m6 activity and leave it at 24°C overnight until the specific activity is 12.0.
Immobilized Tyrosyl-tR showing 00x=7)/ml
NA synthetase (hereinafter referred to as immobilized enzyme) was obtained.

To 2,000 units of this immobilized enzyme or free enzyme, 4 mg of L-chiku 9F1 hydrate, 1) m of disodium adenosine triphosphate
3 mg of g,l,-valine 1 was added, and the pf was adjusted to 95 with a 50 mM sodium carbonate buffer.The total volume was adjusted to 1 ml.The mixture was reacted at 45°C for 2 days.

The amount of tyrosylvalinamide produced was measured using high-performance liquid chromatography on the resulting reaction solution in the same manner as in Example 1. When using the immobilized enzyme.

0.87mg of tyrosylvalinamide was produced,
The amount of tyrosylvalinamide produced when free enzyme was used was 0.30 mg.

(Effect of the invention) According to the present invention. The catalytic ability of aminoacyl-t RNA synthetase is increased, and the reaction rate is significantly improved, especially in a high pH region, so that the yield per unit time can be significantly improved.

Peptide or peptide 83 obtained by the present invention
For example, various hormones and antibiotic peptides such as pradikinin, which has antihypertensive effects, and softstatin, which has endocrine and exocrine suppressive effects.

Useful as other biologically active substances such as taste peptides.

Claims (1)

[Claims] (1) When producing a peptide or peptide derivative by reacting an α-amino acid with an amino acid or an amino acid derivative derived from the amino acid in the presence of an aminoacyl-tRNA synthetase, immobilization is performed on a water-insoluble carrier as an aminoacyl-tRNA synthetase. Aminoacyl-
A method for producing a peptide or a peptide derivative, characterized by using tRNA synthetase.