CN118108581A - Novel diterpenoid compound and preparation method and application thereof - Google Patents
Novel diterpenoid compound and preparation method and application thereof Download PDFInfo
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- CN118108581A CN118108581A CN202410237079.3A CN202410237079A CN118108581A CN 118108581 A CN118108581 A CN 118108581A CN 202410237079 A CN202410237079 A CN 202410237079A CN 118108581 A CN118108581 A CN 118108581A
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Classifications
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
- C07C35/00—Compounds having at least one hydroxy or O-metal group bound to a carbon atom of a ring other than a six-membered aromatic ring
- C07C35/22—Compounds having at least one hydroxy or O-metal group bound to a carbon atom of a ring other than a six-membered aromatic ring polycyclic, at least one hydroxy group bound to a condensed ring system
- C07C35/44—Compounds having at least one hydroxy or O-metal group bound to a carbon atom of a ring other than a six-membered aromatic ring polycyclic, at least one hydroxy group bound to a condensed ring system with a hydroxy group on a condensed ring system having more than three rings
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
- C07C13/00—Cyclic hydrocarbons containing rings other than, or in addition to, six-membered aromatic rings
- C07C13/28—Polycyclic hydrocarbons or acyclic hydrocarbon derivatives thereof
- C07C13/32—Polycyclic hydrocarbons or acyclic hydrocarbon derivatives thereof with condensed rings
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
- C07C13/00—Cyclic hydrocarbons containing rings other than, or in addition to, six-membered aromatic rings
- C07C13/28—Polycyclic hydrocarbons or acyclic hydrocarbon derivatives thereof
- C07C13/32—Polycyclic hydrocarbons or acyclic hydrocarbon derivatives thereof with condensed rings
- C07C13/47—Polycyclic hydrocarbons or acyclic hydrocarbon derivatives thereof with condensed rings with a bicyclo ring system containing ten carbon atoms
- C07C13/52—Azulenes; Completely or partially hydrogenated azulenes
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
- C12N15/09—Recombinant DNA-technology
- C12N15/63—Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
- C12N15/70—Vectors or expression systems specially adapted for E. coli
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
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Abstract
The invention belongs to the technical field of biochemistry and molecular biology, and particularly relates to a novel diterpenoid compound, and a preparation method and application thereof. A novel diterpenoid compound is a compound containing 6-5-6-6 tetracyclic diterpenoid skeleton. The diterpenoid compound provided by the invention has novel structure and function. In addition, the invention constructs the self-sufficient engineering bacteria for producing the novel diterpenoid compounds by utilizing methods of enzyme engineering and genetic engineering. The whole operation process is simple, the process is mature, the cost is low, the whole process does not contain harmful impurities, and the preparation method is nontoxic and is very environment-friendly. The obtained novel diterpenoid compound is expected to be applied to the fields of pharmaceutical chemical industry, agricultural production and the like.
Description
Technical Field
The invention belongs to the technical field of biochemistry and molecular biology, and particularly relates to a novel diterpenoid compound, and a preparation method and application thereof.
Background
Terpene compounds are a class of hydrocarbons containing an integer multiple of isoprene structural units (C 5H8) derived from polyisoprenyl pyrophosphates of limited carbon chain length catalyzed by terpene synthases. Terpenes can be classified into a half terpene (containing 1C 5H8 unit), a monoterpene (containing 2C 5H8 units), a sesquiterpene (containing 3C 5H8 units), a diterpene (containing 4C 5H8 units), and so on, depending on the number of carbon atoms. Terpene compounds and derivatives thereof modified with hetero atoms such as oxygen, nitrogen, halogen, etc. are collectively called terpenoids. They are the most numerous natural products in nature, and more than one hundred thousand terpenoids have been reported. The terpenoid is used as primary or secondary metabolite in human, animal, plant and microorganism, has various structure types and wide distribution, and can be biosynthesized in more than 14000 organisms in 1109 families in nature. Meanwhile, the terpenoid has rich biological functions such as aromatic smell/flavor, sterilization, anti-tumor, antiviral, anti-inflammatory, immunosuppression, regulation of physiological metabolic processes in human and animal bodies and the like, and has important application value and potential in the industries of medicines, foods, essence and perfume. Representative terpenoids, such as the sesquiterpene lactone derivative artemisinin isolated from Artemisia annua taught by Nobel prize Tu Youyou, are not only anticancer and anti-inflammatory, but also antimalarial special effect drugs saving tens of millions of lives; paclitaxel isolated from taxus genus plants was successfully applied to the treatment of ovarian cancer and breast cancer clinically; tanshinone extracted from traditional Chinese medicine salvia miltiorrhiza has the functions of resisting bacteria, diminishing inflammation, promoting blood circulation, removing blood stasis and the like, and is widely applied to the treatment of cardiovascular and cerebrovascular diseases; steroid hormones, which are indispensable to humans and mammals, are widely involved in the regulation of growth and metabolism in vivo. In order to meet urgent demands of people on healthy life, the excavation of novel active terpenoids or terpene skeletons is always an important link in the development of new drugs.
However, due to resource and technical limitations, especially the fact that plants are used as main sources of terpenoids, the disadvantages of long growth cycle, low yield of terpenoids and high acquisition cost exist, so that researchers are more and more difficult to directly excavate terpenoids from the nature. Moreover, the novel terpenoid obtained by the chemical synthesis strategy has the defects of more reaction steps, harsh conditions, poor catalytic selectivity, environmental pollution and the like, and more importantly, the novel terpenoid has limited capability of obtaining a brand new structural framework. In recent years, with the rapid development of structural biology and enzyme engineering, by analyzing the crystal structure and catalytic mechanism of terpene synthases, the important amino acid sites of the terpene synthases are replaced by adopting a site-directed mutation technology so as to change the stability of a hydrogen bond network or a carbocation intermediate of an active pocket of the terpene synthases, thereby realizing the biosynthesis of novel terpene compounds, and the novel terpene compounds have become an important means for excavating novel terpene compounds for basic and application science research.
Diterpene synthase VenA from Streptomyces venezuelans (Streptomyces venezuelae ATCC 15439) catalyzes the biosynthesis of the new backbone diterpene venezuelan A containing 5-5-6-7 tetracyclic groups from geranylgeranyl pyrophosphate (GERANYLGERANYL PYROPHOSPHATE, GGPP). VenA has good expression in Escherichia coli, good catalytic efficiency and good substrate broad property, and is expected to develop into bacterial diterpene synthase model enzyme. However, the search found that there is no report on the production of novel diterpenoids venezuelaenol A, venezuelaenol B, (1S, 3E,7E,11R, 12S) -3,7,18-dolabellatriene and DICTYTRIENE C by engineering diterpenoid synthase VenA or the production of corresponding venezuelaenol A, venezuelaenol B, (1S, 3E,7E,11R, 12S) -3,7,18-dolabellatriene and DICTYTRIENE C by expressing VenA mutants in E.coli by heterologous expression methods at home and abroad, and isolation and purification thereof.
Disclosure of Invention
Aiming at the defects of the prior art, the invention aims to provide a novel diterpenoid compound, and a preparation method and application thereof.
In order to achieve the purpose of the invention, the technical scheme adopted by the invention is as follows: a novel diterpenoid compound is a compound containing a 6-5-6-6 tetracyclic diterpenoid skeleton, and the structural formula of the novel diterpenoid compound is shown as the following formula:
The invention also provides a preparation method of the novel diterpenoid compound, and a venA mutant gene is expressed in escherichia coli through a heterologous expression strategy to obtain mutant protein; the mutant protein is separated and purified to obtain the compound.
The invention also provides application of the novel diterpenoid compounds in pharmaceutical chemical industry and agricultural production, wherein the application in the pharmaceutical chemical industry does not comprise a disease diagnosis and treatment method.
Compared with the prior art, the invention has the following beneficial effects: the invention provides a novel diterpenoid compound, and a preparation method and application thereof. The inventor modifies key amino acids in a diterpenoid synthase VenA active pocket from S.venezuelae ATCC 15439 in a point mutation mode to obtain mutant genes, and realizes the expression of diterpenoid compounds in escherichia coli. Finally, 4 novel diterpenoid compounds are obtained through separation and purification, and the diterpenoid compounds have novel structures and functions. In addition, the innovation point of the invention is also embodied in the construction of self-sufficient engineering bacteria for producing novel diterpenoid compounds by utilizing methods of enzyme engineering and genetic engineering. The whole operation process is simple, the process is mature, the cost is low, the whole process does not contain harmful impurities, and the preparation method is nontoxic and is very environment-friendly. The obtained novel diterpenoid compound is expected to be applied to the fields of pharmaceutical chemical industry, agricultural production and the like.
Drawings
FIG. 1 is an ellipsoidal diagram of a three-dimensional structure of a compound according to an embodiment of the present invention;
FIG. 2 is a diagram of a gas mass spectrum of venezuelaenol A provided by an embodiment of the present invention;
FIG. 3 shows key correlation signals of 1H-1 H COSY, HMBC, and NOESY of venezuelaenol A provided by an embodiment of the present invention;
FIG. 4 is a 1 H NMR chart of venezuelaenol A in a CD 3 CN provided by an embodiment of the invention;
FIG. 5 is a 13 C NMR chart of venezuelaenol A in a CD 3 CN provided by an embodiment of the invention;
FIG. 6 is a diagram of HSQC of venezuelaenol A in CD 3 CN according to an embodiment of the present invention;
Fig. 7 is a HMBC diagram of venezuelaenol A in a CD 3 CN provided by an embodiment of the present invention;
FIG. 8 is a diagram of a 1H-1 H COSY of venezuelaenol A in a CD 3 CN provided by an embodiment of the present invention;
FIG. 9 is a NOESY diagram venezuelaenol A in a CD 3 CN provided by an embodiment of the present invention;
FIG. 10 is a diagram of a gas mass spectrum of venezuelaenol B provided by an embodiment of the present invention;
FIG. 11 shows key correlation signals of 1H-1 H COSY, HMBC, and NOESY of venezuelaenol B provided by an embodiment of the present invention;
FIG. 12 is a 1 H NMR chart of venezuelaenol B in a CD 3 CN provided by an embodiment of the invention;
FIG. 13 is a 13 C NMR chart of venezuelaenol B in a CD 3 CN provided by an embodiment of the invention;
FIG. 14 is a diagram of a 13 C-DEPT 135℃in a CD 3 CN venezuelaenol B provided by an embodiment of the present invention;
FIG. 15 is a diagram showing HSQC of venezuelaenol B in CD 3 CN according to an embodiment of the present invention;
Fig. 16 is a HMBC diagram of venezuelaenol B in a CD 3 CN provided by an embodiment of the present invention;
FIG. 17 is a diagram of a 1H-1 H COSY of venezuelaenol B in a CD 3 CN provided by an embodiment of the present invention;
Fig. 18 is a view of venezuelaenol B NOESY in CD 3 CN, provided in an embodiment of the present invention.
Detailed Description
The present invention will be described in detail with reference to specific examples. The following examples are only preferred embodiments of the present invention, and are not intended to limit the present invention in any way, and any simple modification, equivalent changes and modification made to the embodiments according to the technical substance of the present invention are within the scope of the technical solution of the present invention.
The basic molecular biology experimental techniques such as PCR amplification, plasmid extraction, transformation, etc., employed in the examples of the present invention, unless otherwise specified, are usually performed according to conventional methods, and can be specifically described in "molecular cloning Experimental guidelines" (third edition) (Sambrook J, russell DW, janssen K, ARGENTINE J Huang Peitang, et al, 2002, beijing: scientific press), or according to the specifications provided by the relevant manufacturers.
Plasmids pET28b-venA and pET28b-venD used in the examples of the invention were stored in the laboratory; ESCHERICHIA COLI BL21 (DE 3) competent cells were purchased from Qingdao Optimaceae. The construction method of the engineering bacterium Eco-P (ESCHERICHIA COLI BL (DE 3)/pACYC-mavEmavS & pTrc-low) of the escherichia coli is Microb Cell Fact 15,74 (2016).
The one-step cloning kit used in the examples of the present invention was purchased from the company Nanjinouzan; agarose gel DNA recovery kit was purchased from Omega company; coli expression vector pET28b was purchased from Invitrogen; high fidelity DNA polymerase was purchased from Takara; restriction enzymes were purchased from sirolimus; PCR primer synthesis and DNA sequencing were done by qinghao.
The seed culture medium of the enterobacteria related to the embodiment of the invention is LB liquid culture medium, and the formula is as follows: 10g/L of tryptone, 5g/L of yeast extract and 10g/L of NaCl; the fermentation medium is a TB medium, and the formula is as follows: 12g/L tryptone, 24g/L yeast extract, 40g/L glycerol and K 2HPO4 9.4g/L,KH2PO4 2.2.2 g/L.
Example 1 Gene expression vector construction
The linear expression vectors of diterpene synthase VenA mutant coding genes venA W107A、venAV111W and venA F185A are obtained by respectively amplifying with primers W107A-F/W107A-R, V111W-F/V111W-R and F185A-F/F185A-R (Table 1) by taking pET28b-venA as a template, and the reaction system is as follows: 5X PRIMESTAR GXL BUFFER. Mu.L, 200. Mu.M dNTPs, 4. Mu.L DMSO, 0.3. Mu.M each of the upstream and downstream primers, an appropriate amount of DNA template (10-100 ng), 2.5U of high-fidelity polymerase (PRIMESTAR GXL DNA polymerase), and a ddH 2 O make-up to 50. Mu.L; the reaction conditions are as follows: pre-denaturation at 98℃for 5min, denaturation at 98℃for 30s, annealing at 60℃for 15s, extension at 68℃for 1kb/min, set time according to the fragment length of interest, 35 cycles of reaction, and finally extension at 68℃for 10min. The amplified linearized plasmid pET28b-venA W107A、pET28b-venAV111A、pET28b-venAF185A is purified by a nucleic acid purification kit, and then template DNA is further removed by DpnI digestion.
In this example, the nucleotide sequences of mutant encoding genes venA F185A、venAV111W and venA W107A are shown in SEQ ID NOs: 1. SEQ ID NO: 3. SEQ ID NO:5, the amino acid sequences of the encoded mutant proteins are respectively shown in SEQ ID NO: 2. SEQ ID NO: 4. SEQ ID NO: shown at 6. In other embodiments of the invention, the sequence of the mutant encoding gene may also be a sequence encoding a sequence as set forth in SEQ ID NO: 2. SEQ ID NO: 4. SEQ ID NO:6 or a nucleotide sequence corresponding to an amino acid sequence having a sequence identity of more than 80% to the amino acid sequence described above.
Subsequently, PCR was performed using the vector pET28b-venD expressing geranylgeranyl pyrophosphate synthase as a template and using the primer T7VenD-F/T7VenD-R to obtain a DNA fragment containing the T7 promoter and terminator sequences and venD, and the template DNA was removed by DpnI digestion. Finally VenD was cloned into the above prepared linearized pET28b-venA W107A、pET28b-venAV111W and pET28b-venA F185A to give co-expression vectors pET28b-venA W107AD、pET28b-venAV111W D and pET28b-venA F185A D containing the venA mutant gene of 2T 7 promoters and venD.
Table 1: primer sequence for amplification of the Gene encoding diterpene synthase VenA mutant obtained in example 1
EXAMPLE 2 construction of recombinant E.coli, fermentation production of novel diterpenoid Compounds
And (3) converting pET28b-venA W107AD、pET28b-venAV111W D and pET28b-venA F185A D into E.coli engineering bacteria Eco-P (ESCHERICHIA COLI BL (DE 3)/pACYC-mavEmavS & pTrc-low) chemically competent which are prepared in advance and high in yield of dimethyl propenyl pyrophosphoric acid and isopentenyl pyrophosphoric acid, so as to obtain recombinant E.coli engineering bacteria Eco-P/pET28b-venA W107AD、Eco-P/pET28b-venAV111W D and Eco-P/pET28b-venA F185A D for self-sufficient production of novel diterpenoid compounds.
The Eco-P/pET28b-venA F185A D was inoculated from LB solid plates (tryptone 10g/L, yeast extract 5g/L, naCl 10g/L,15g/L agar powder) into 100mL of LB liquid medium (tryptone 10g/L, yeast extract 5g/L, naCl 10 g/L) containing 50. Mu.g/mL kanamycin, 25. Mu.g/mL chloramphenicol and 100. Mu.g/mL ampicillin, and cultured overnight at 37℃and 220 rpm. Then, the seed solution was transferred to 10L of TB medium (tryptone 12g/L, yeast extract 24g/L, glycerol 40g/L, K 2HPO4 9.4g/L,KH2PO4 2.2.2 g/L) containing 50. Mu.g/mL kanamycin, 25. Mu.g/mL chloramphenicol, and 100. Mu.g/mL ampicillin in a volume ratio of 1:100, and cultured at 37℃and 220 rpm. When OD 600 reached 1.0-1.5, isopropyl- β -D-thiogalactoside was added at a final concentration of 0.2mM, protein expression was induced and biosynthesis venezuelaenol A and venezuelaenol B was performed at 18℃and 220 rpm.
Example 3 isolation and purification and Structure identification of novel diterpene Compounds
After the recombinant escherichia coli Eco-P/pET28b-venA W107AD、Eco-P/pET28b-venAV111W D and Eco-P/pET28b-venA F185A D are fermented for 3 days, adding an equal volume of ethyl acetate for extraction for three times, and evaporating the extract by using a rotary evaporator to obtain an extract. Then, the extract was extracted four times with 50-100mL of n-hexane to remove the polar components. Finally, the n-hexane extract was evaporated to dryness and dissolved in acetonitrile for semi-preparation of the diterpene compound by reversed phase C18 chromatographic column.
A specific column was YMC-18 column (10X 250mm,5 μm) and the elution procedure for preparation venezuelaenol A and venezuelaenol B was: 100% acetonitrile at a flow rate of 1.5mL/min (0-17 min); the flow rate is 2.0mL/min (7.5-23 min); the flow rate is 2.5mL/min (23.5-29 min); the flow rate was 3mL/min (29.5-35 min). The elution procedure for preparation DICTYTRIENE C and (1S, 3E,7E,11R, 12S) -3,7,18-dolabellatriene was: 100% acetonitrile was eluted for 30min at a flow rate of 3mL/min.
The venezuelaenol A finally obtained is colorless transparent oil-like after being dried by nitrogen, and the specific rotation [ alpha ] 2 D 0＝-51.1(c 0.083,CH3 CN). Further, in order to complete the structure identification and analysis of venezuelaenol A, 2mg of the compound was dissolved in 0.2mL of chromatographically pure acetonitrile, and naturally volatilized at 4℃for about one week, needle-like crystals were formed for X-ray single crystal diffraction analysis to obtain the three-dimensional structure thereof, as shown in FIG. 1.
The chemical structure of the compound venezuelaenol A (molecular formula: C 20H34 O) is shown as a formula (1):
the chemical shifts were assigned to table 2 by gas phase mass spectrometry as shown in fig. 2 and by nuclear magnetic resonance spectroscopy as shown in fig. 3-9.
Table 2: 1H NMR(600MHz,CD3 CN) and 13C NMR(151MHz,CD3 CN) chemical shift assignment of venezuelaenol A in examples
The venezuelaenol B obtained finally is dried by nitrogen and is white powder, and the specific rotation [ alpha ] 2 D 0＝-24.6(c 0.133,CH3 CN). From the gas phase mass spectrum (FIG. 10) and the nuclear magnetic resonance spectrum analysis (FIGS. 11-18), diterpene compounds (molecular formula: C 20H34 O) containing a new skeleton of 6-5-6-6 were confirmed and chemical shifts thereof were attributed to Table 3.
The chemical structural formula of the compound venezuelaenol B is shown as a formula (2):
Table 3: 1H NMR(600MHz,CD3 CN) and 13C NMR(151MHz,CD3 CN) chemical shift assignment of venezuelaenol B in examples
Example 4 evaluation of Compounds venezuelaenol A, venezuelaenol B for use in medicine and agriculture
To investigate the antibacterial activity of compounds venezuelaenol A, venezuelaenol B, the novel diterpenoid compounds were assayed against seven gram-positive strains (ampicillin-resistant Bacillus subtilis (AMPICILLIN-RESISTANT BACILLUS SUBTILIS), bacillus subtilis RIK1285 (B. Subtilis RIK 1285), bacillus subtilis 168 (B. Subtilis 168), bacillus cereus (Bacillus cereus), potato rot germ (Clavibacter michiganense subsp. Sepedonicus), staphylococcus aureus (Staphylococcus aureus ATCC 25923) and pubic symphysis) according to a micro broth dilution method using kanamycin as a positive control
Mycobacterium smegmatis (Mycolicibacterium SMEGMATIS MC)) and four gram-negative plant pathogens (pectobacterium carotovorum (Pectobacterium carotovorum subsp. Carotovorum), brevibacterium cucumeris (Pseudomonas syringae pv. Lachrymans), xanthomonas campestris (Xanthomonas campestris pv. Campestris) and Brevibacterium oryzae (Xanthomonas oryzae pv. Oryzicola RS 105)).
The above compound was dissolved in dimethyl sulfoxide to prepare a mother liquor with a concentration of 256. Mu.g/mL, and the mother liquor was added to LB medium containing 10 6 cfu/mL of indicator bacteria to be diluted stepwise to a concentration of 256, 128, 64, 32, 16, 8, 4, 2, 1, 0.5, 0.25, 0.125. Mu.g/mL, etc., and only an equal volume of dimethyl sulfoxide was added to the negative control. After the indicator bacteria are continuously cultured for 24 hours at 28 ℃ and 200rpm, observing that the concentration of the compound corresponding to the clarified sample hole is the minimum inhibitory concentration (Minimum Inhibitory Concentration, MIC). As shown in table 6, compound venezuelaenol A exhibited moderate inhibitory activity against bacillus subtilis 168, staphylococcus aureus ATCC 25923, and cucumber bacterial angular leaf spot bacteria; wherein, the antibacterial spectrum of the compound DICTYTRIENE C is wider, and the compound shows the inhibitory activity on 9 indicator bacteria to be detected and the like. The compound venezuelaenol A has great application potential in the fields of medicine and agriculture.
Although compound venezuelaenol B did not exhibit significant bacteriostatic activity against the indicator bacteria. However, since many active natural products are reported to be modified by a dolastane skeleton and venezuelaenol B contains a complex ring system similar to venezuelaenol A, the compounds venezuelaenol B and (1S, 3E,7E,11R, 12S) -3,7,18-dolabellatriene are expected to be used as precursors to be further modified by oxidation, glycosylation and the like to obtain corresponding active lead molecules, and the active lead molecules are applied to the fields of medicines, agricultural production and the like.
Table 6: examples the invention provides novel diterpene compounds useful in the antibacterial activity test
Claims (3)
1. A novel diterpenoid compound, which is characterized in that: the compound is a compound containing a 6-5-6-6 tetracyclic diterpenoid skeleton, and has the structural formula:
。
2. The method for preparing the novel diterpenoid compound according to claim 1, which is characterized in that: expressing venA mutant genes in escherichia coli through a heterologous expression strategy to obtain mutant proteins; the mutant protein is separated and purified to obtain the compound.
3. The use of a novel diterpenoid compound according to claim 1 in pharmaceutical chemical or agricultural production, characterized in that: the application in pharmaceutical and chemical industry does not include methods for diagnosing and treating diseases.
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