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Substrate selection of adenylation domains for nonribosomal peptide synthetase (NRPS) in bacillamide C biosynthesis by marine Bacillus atrophaeus C89

  • Biotechnology Methods - Original Paper
  • Published:
Journal of Industrial Microbiology & Biotechnology

Abstract

Nonribosomal peptide synthetases (NRPSs) are multi-modular enzymes involved in the biosynthesis of natural products. Bacillamide C was synthesized by Bacillus atrophaeus C89. A nonribosomal peptide synthetase (NRPS) cluster found in the genome of B. atrophaeus C89 was hypothesized to be responsible for the biosynthesis of bacillamide C using alanine and cysteine as substrates. Here, the structure analysis of adenylation domains based on homologous proteins with known crystal structures indicated locations of the substrate-binding pockets. Molecular docking suggested alanine and cysteine as the potential substrates for the two adenylation domains in the NRPS cluster. Furthermore, biochemical characterization of the purified recombinant adenylation domains proved that alanine and cysteine were the optimum substrates for the two adenylation domains. The results provided the in vitro evidence for the hypothesis that the two adenylation domains in the NRPS of B. atrophaeus C89 preferentially select alanine and cysteine, respectively, as a substrate to synthesize bacillamide C. Furthermore, this study on substrates selectivity of adenylation domains provided basis for rational design of bacillamide analogs.

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Acknowledgements

This work was supported by the National Natural Science Foundation of China (NSFC) (31300104).

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Author notes

  1. Fengli Zhang and Yukun Wang are contributed equally to this work.

Authors and Affiliations

  1. Marine Biotechnology Laboratory, State Key Laboratory of Microbial Metabolism and School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai, China

    Fengli Zhang, Yukun Wang, Qun Jiang, Loganathan Karthik & Zhiyong Li

  2. Computational Chemistry and Molecular Bioinformatics Laboratory, State Key Laboratory of Microbial Metabolism and School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai, China

    Qihua Chen & Yi-Lei Zhao

Authors
  1. Fengli Zhang
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  2. Yukun Wang
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  4. Qihua Chen
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  7. Zhiyong Li
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Corresponding author

Correspondence to Zhiyong Li.

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Fig. S1

The structures of bacillamide A, bacillamide B, bacillamide C, bacillamide D and bacillamide E

Fig. S2

NRPSpredictor2 prediction report for the A1 and A2 domains. Location of the A domain within the sequence and the bit score of the PFAM-HMM are provided. The green checkmark signals indicate that the signature sequence lies within the applicability domain. The list of predictions is provided along with the scores of the respective SVM predictors. The final row provides the nearest sequence neighbor in the NRPSpredictor2 database (based on the Stachelhaus code) and the respective sequence identity (%)

Fig. S3

Ramachandran plot atlas of the A-domain structure model (a) and Ramachandran plot atlas of the A-domain structure model after energy minimization (b). (1) A1 domain; (2) A2 domain. The figure is plotted using Discovery Studio

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Zhang, F., Wang, Y., Jiang, Q. et al. Substrate selection of adenylation domains for nonribosomal peptide synthetase (NRPS) in bacillamide C biosynthesis by marine Bacillus atrophaeus C89. J Ind Microbiol Biotechnol 45, 335–344 (2018). https://doi.org/10.1007/s10295-018-2028-2

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  • DOI: https://doi.org/10.1007/s10295-018-2028-2

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