Complete genome sequence of high-yield strain S. lincolnensis B48 and identification of crucial mutations contributing to lincomycin overproduction

Ruida Wang; Fanjing Kong; Haizhen Wu; Bingbing Hou; Yajing Kang; Yuan Cao; Shiwei Duan; Jiang Ye; Huizhan Zhang

Synthetic and Systems Biotechnology (Jun 2020)

Complete genome sequence of high-yield strain S. lincolnensis B48 and identification of crucial mutations contributing to lincomycin overproduction

Ruida Wang,
Fanjing Kong,
Haizhen Wu,
Bingbing Hou,
Yajing Kang,
Yuan Cao,
Shiwei Duan,
Jiang Ye,
Huizhan Zhang

Affiliations

Ruida Wang: State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai, 200237, China
Fanjing Kong: State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai, 200237, China
Haizhen Wu: State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai, 200237, China; Department of Applied Biology, East China University of Science and Technology, Shanghai, 200237, China
Bingbing Hou: State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai, 200237, China; Department of Applied Biology, East China University of Science and Technology, Shanghai, 200237, China
Yajing Kang: State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai, 200237, China
Yuan Cao: State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai, 200237, China
Shiwei Duan: Medical Genetics Center, School of Medicine, Ningbo University, Ningbo, 315211, China
Jiang Ye: State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai, 200237, China; Department of Applied Biology, East China University of Science and Technology, Shanghai, 200237, China; Corresponding author. State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai, 200237, China.
Huizhan Zhang: State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai, 200237, China; Department of Applied Biology, East China University of Science and Technology, Shanghai, 200237, China; Corresponding author. State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai, 200237, China.

Journal volume & issue: Vol. 5, no. 2
pp. 37 – 48

Abstract

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The lincosamide family antibiotic lincomycin is a widely used antibacterial pharmaceutical generated by Streptomyces lincolnensis, and the high-yield strain B48 produces 2.5 g/L lincomycin, approximately 30-fold as the wild-type strain NRRL 2936. Here, the genome of S. lincolnensis B48 was completely sequenced, revealing a ~10.0 Mb single chromosome with 71.03% G + C content. Based on the genomic information, lincomycin-related primary metabolism network was constructed and the secondary metabolic potential was analyzed. In order to dissect the overproduction mechanism, a comparative genomic analysis with NRRL 2936 was performed. Three large deletions (LDI-III), one large inverted duplication (LID), one long inversion and 80 small variations (including 50 single nucleotide variations, 13 insertions and 17 deletions) were found in B48 genome. Then several crucial mutants contributing to higher production phenotype were validated. Deleting of a MarR-type regulator-encoding gene slinc377 from LDI, and the whole 24.7 kb LDII in NRRL 2936 enhanced lincomycin titer by 244% and 284%, respectively. Besides, lincomycin production of NRRL 2936 was increased to 7.7-fold when a 71 kb supercluster BGC33 from LDIII was eliminated. As for the duplication region, overexpression of the cluster situated genes lmbB2 and lmbU, as well as two novel transcriptional regulator-encoding genes (slinc191 and slinc348) elevated lincomycin titer by 77%, 75%, 114% and 702%, respectively. Furthermore, three negative correlation genes (slinc6156, slinc4481 and slinc6011) on lincomycin biosynthesis, participating in regulation were found out. And surprisingly, inactivation of RNase J-encoding gene slinc6156 and TPR (tetratricopeptide repeat) domain-containing protein-encoding gene slinc4481 achieved lincomycin titer equivalent to 83% and 68% of B48, respectively, to 22.4 and 18.4-fold compared to NRRL 2936. Therefore, the comparative genomics approach combined with confirmatory experiments identified that large fragment deletion, long sequence duplication, along with several mutations of genes, especially regulator genes, are crucial for lincomycin overproduction.

Published in Synthetic and Systems Biotechnology

ISSN: 2405-805X (Online)
Publisher: KeAi Communications Co., Ltd.
Country of publisher: China
LCC subjects: Technology: Chemical technology: Biotechnology; Science: Biology (General)
Website: https://www.keaipublishing.com/en/journals/synthetic-and-systems-biotechnology/

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