Screening and functional validation of endogenous promoters from Bacillus amyloliquefaciens for fengycin synthesis

doi:10.14088/j.cnki.issn0439-8114.2026.02.031

Abstract

Abstract: Using Bacillus amyloliquefaciens TF28 as the host, and by integrating genomic and transcriptomic data, 12 upstream endogenous promoters from highly expressed genes were screened. By constructing a GFP reporter system and evaluating the activity of each promoter in Bacillus, it was found that the promoter strengths of PspoVG, PqoxC, PqoxB, and Pgpd were higher than that of the commonly used promoter P43. When driven by the PspoVG promoter, the transcriptional level of the fenC gene was 4.62 times that of the P43 promoter, and the fengycin yield increased from 84.3 mg/L to 101.2 mg/L. Regarding promoter structure, the sequences of the -35 and -10 regions of PspoVG were highly consistent with the consensus sequences of typical strong promoters, which might be the structural basis for its high-efficiency transcriptional drive.

Key words: fengycin, Bacillus amyloliquefaciens, endogenous promoter, screening, functional validation

CLC Number:

Q933

ZHOU Lu, TIAN Yuan, YU Chong, XIA Hai-hua, YAN Geng-xuan. Screening and functional validation of endogenous promoters from Bacillus amyloliquefaciens for fengycin synthesis[J]. HUBEI AGRICULTURAL SCIENCES, 2026, 65(2): 209-214.

References

[1] CAO G, LIU Y, LIU G, et al.A novel strong promoter of Bacillus amyloliquefaciens and its application in enhancing fengycin production[J].Applied microbiology and biotechnology, 2020, 104(15): 6671-6682.
[2] CHEN X, LIU Y, WANG J, et al.Enhancing fengycin production in Bacillus velezensis by promoter engineering[J].Journal of industrial microbiology & biotechnology, 2019, 46(5): 719-730.
[3] DUNN A K, HANDELSMAN J.A vector for promoter trapping in Bacillus cereus[J].Gene, 2021, 768: 145263.
[4] FAN B, WANG C, SONG X, et al.Bacillus velezensis FZB42 in plant growth-promotion and molecular mechanisms of its broad-spectrum antimicrobial activity[J].Trends in microbiology, 2018, 26(7): 603-615.
[5] GU Y, XU X, WU Y, et al.Advances in Bacillus lipopeptides: structures, biosynthesis and properties[J].Applied microbiology and biotechnology, 2017, 101(20): 7567-7576.
[6] HU F, LIU Y, LI S, et al.A novel promoter screening system in Bacillus subtilis based on green fluorescent protein[J].Journal of microbiological methods, 2018, 145: 1-7.
[7] JIANG J,GAO L,BIE X,et al.Identification of a strong promoter for the heterologous expression of lipopeptides in Bacillus subtilis[J].Biotechnology letters, 2019, 41(4-5): 549-557.
[8] KIM P I, BAI H, BAI D, et al.Purification and characterization of a lipopeptide produced by Bacillus thuringiensis CMB26[J].Journal of applied microbiology, 2011, 111(4): 890-899.
[9] LI Y, LI Z, LIU H, et al.A strong promoter from Bacillus amyloliquefaciens promotes the expression of heterologous proteins[J].Protein expression and purification, 2020, 166: 105507.
[10] LIU J, LIU Y, WANG Y, et al.A high-throughput screening method for strong promoters in Bacillus subtilis[J].Scientific reports, 2019, 9(1): 11806.
[11] LUO C, LIU X, ZHOU H, et al.Promoter engineering for the synthesis of lipopeptides in Bacillus amyloliquefaciens[J].Metabolic engineering, 2022, 69: 1-10.
[12] MA Z, HU J, WANG X, et al.A novel promoter from Bacillus velezensis for the expression of antifungal lipopeptides[J].Microbial cell factories, 2020, 19(1): 1-12.
[13] PANG X, LIN X, WANG J, et al.Identification and characterization of a strong promoter Pg3 from Bacillus subtilis[J].Journal of biotechnology, 2018, 267: 1-8.
[14] QIU Y, ZHANG J, LI L, et al.Engineering Bacillus subtilis for the production of fengycin analogues[J].ACS synthetic biology, 2020, 9(11): 3076-3086.
[15] RAUTENBACH M,TROSTYANSKAJA I B,VIJLOEN A,et al.The role of fengycin in the biological control of Fusarium species[J].Journal of applied microbiology, 2016, 120(5): 1254-1267.
[16] SUN H, BIE X, LU F, et al.Enhancement of surfactin production in Bacillus amyloliquefaciens by promoter replacement[J].Applied microbiology and biotechnology, 2019, 103(21-22): 9097-9106.
[17] WANG J, LIU J, WANG X, et al.A high‐efficiency CRISPR/Cas9 system for genome editing in Bacillus amyloliquefaciens[J].Journal of basic microbiology, 2021, 61(2): 105-115.
[18] WANG L, LIU Y, LIU J, et al.A novel promoter Pfen for the expression of fengycin synthetase in Bacillus subtilis[J].Enzyme and microbial technology, 2018, 110: 1-8.
[19] XU D, CHEN X, LIU Y, et al.A strong constitutive promoter from Bacillus amyloliquefaciens and its application in lipopeptide production[J].Biotechnology and bioengineering, 2020, 117(6): 1781-1791.
[20] YANG H, LI X, LIU Y, et al.A novel promoter screening system in Bacillus subtilis using a secreted luciferase reporter[J].Journal of microbiological methods, 2017, 142: 1-7.
[21] ZHANG L, SUN C.Fengycins, cyclic lipopeptides from marine Bacillus subtilis strains, kill the plant-pathogenic fungus Magnaporthe grisea by inducing reactive oxygen species production and chromatin condensation[J].Applied and environmental microbiology, 2018, 84(18): e00445-18.
[22] ZHANG Y, LIU Y, LIU J, et al.A novel strong promoter Pqox from Bacillus amyloliquefaciens and its application in the production of antifungal lipopeptides[J].Journal of agricultural and food chemistry, 2021, 69(12): 3675-3683.
[23] ZHAO X, KUIPERS O P.Identification and classification of known and novel antimicrobial compounds produced by Bacillusspp. isolated from different environments[J].Microbial biotechnology, 2021, 14(3): 1073-1100.