Penicilliumsp.MYA5中microperfuranone聚酮合酶基因在構(gòu)巢曲霉中的異源表達(dá)

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中圖分類號：Q786 文獻(xiàn)標(biāo)志碼：A

Heterologous expression of the microperfuranone PKS from Penicillium sp.MYA5 in Aspergillus nidulans

MengXianchao1，HuBo2，YuHaobing2，SunYuanyuan2，andLiuXiaoyu2 （1 SchoolofOceanFoodandBilogicalEngieeingJangsuOceanUnversityLianungang2O5;2LANavalMedicalCeter Naval Medical University， Shanghai 200433）

Abstract Objective To enhance the production of the antimicrobial compound microperfuranone derived from the polar endophytic fungus Penicillium sp.MYA5 through gene mining and heterologous expression strategies， and to evaluate its bioactivity.MethodsGenome sequencing data for Penicillium sp.MYA5 were analyzed using antiSMASH to identify biosynthetic gene clusters.The polyketide synthase gene mfpksA wascloned and inserted into the recombinant plasmid pYTU-mfpksA.Screening positive clones through yeastand Escherichia coli transformation. The recombinant plasmid was introduced into Aspergillus nidulans LO8030，and screened on CHsorbitol plates to obtain therecombinant strainThe plasmid was transformed into Saccharomyces cerevisiaeBJ5464andEscherichia coli TOP10 forvalidation，heterologous expression was performed in Aspergilus nidulans LO8030，followed by fermentation， HPLC analysis，and structural elucidation via NMR and massspectrometry. Antimicrobial activity was assessed against pathogenic strains using the broth microdilution method. ResultsThe target compound microperfuranone was successfullyisolated from the heterologous expression strain，achieving anapproximately3.88- fold increase in yield compared to the MYA5 strain.Antimicrobial activity assays revealed that microperfuranone exhibited a minimum inhibitory concentration （MIC） of 16μg/mL against both Fusarium oxysporum and Vibrio harveyi. Notably，its inhibitory activity against F. oxysporum was significantly superior to chloramphenicol. Conclusion The heterologous expression strategy successfully enhanced the production of microperfuranone， confirming the effectiveness of biosynthetic gene cluster mining and heterologous expression in natural product discovery.The compound exhibited significant inhibitory activity against plant pathogens，providing a theoretical foundation for the antimicrobial application of polar microbial resources.

Key WordsMicroperfuranone; Penicillium sp.; PKS; Heterologous expression; Polyketide compounds

呋喃酮類化合物是一類含有氧原子的五元雜環(huán)芳香化合物，具有抗菌、抗氧化、抗炎和抗癌等多種生物活性[1-3]，已開發(fā)上市的藥物有痢特靈、呋喃酮唑片等，真菌中已報道的呋喃酮類化合物多數(shù)都是通過聚酮合成酶（PKS）途徑合成的，如asperfuranone，gregatinA等[4-5]，其不同的官能團(tuán)具有不同的生物活性[]，microperfuranone是1個聯(lián)苯呋喃酮化合物，其結(jié)構(gòu)為5-羥基-4-芐基-3-苯基-2（5H）-呋喃酮，已從真菌Aspergilluscarneus，Aspergillussp.Y-12和AcremoniumpilosumF47等菌株中分離得到[7-10]，具有抗菌，抗病毒等生物活性[1]，但通過常規(guī)分離手段的產(chǎn)量相對較低，可以采取基因工程技術(shù)對其生物合成基因進(jìn)行異源表達(dá)，不僅能避免原始宿主復(fù)雜代謝網(wǎng)絡(luò)對產(chǎn)物合成的干擾，而且目標(biāo)化合物的產(chǎn)量也會有所提高。（剩余12457字）