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type Journal Article
authors Wu, L.; Liu, A.; Ma, C.; Han, F.; Zhang, H.; Gao, Q.; Wang, Y.
title Cloning of 3-amino-5-hydroxybenzoic acid (AHBA) synthase gene from Streptomyces sp. 4353 and its expression in E. coli
journal Chin. J. Antibiot.
Activity 4.2.1.144
Family 4.2.1.144
sel unselected
ui notinPubmedyet
year (2009)
volume 34
pages 24-29
 
abstract Objective Cloning of 3-amino-5-hydroxybenzoic acid (AHBA) synthase gene from Streptomyces sp. 4353 and its expression in E. coli. Method Five genes, i. e. genes for amino DHQ dehydratase, aminoquinate/shikimate dehydrogenase, AHBA synthase, oxidoreductase and phosphatase, all involved in AHBA biosynthesis, are usually genetically linked as an AHBA biosynthetic gene cluster. Primers were designed to amplify the upstream and downstream DNA sequences of a known 737bp DNA encoding the middle conserved region of the AHBA synthase gene from Streptomyces sp. 4353 by PCR. The PCR-amplified DNA fragments were cloned into pMD18-T, then sequenced. The AHBA synthase gene thus obtained from Streptomyces sp. 4353 was expressed in BL21-Codon Plus(DE3)-RIL using the E. coli expression vector pET24a( + ), and detected by SDS-PAGE. Results A 2, 872bp DNA fragment from Streptomyces sp. 4353 was assembled and deposited in NCBI GenBank under the Accession No. EU734184. Frame-plot and Blast analysis indicated that it contained the AHBA synthase gene, the oxidoreductase gene and the 5' region of the phosphatase gene for AHBA biosynthesis. Protein bands of about 42. 7ku, the expected molecular mass of AHBA synthase were visualized clearly on the SDS-PAGE, indicating that the AHBA synthase gene from Streptomyces sp. 4353 was expressed successfully in BL21-Codon Plus(DE3)-RIL. Conclusion The cloning of the AHBA synthase gene from Streptomyces sp. 4353 and its expression in E. coli provided the basis for its further functional characterization. The 2, 872bp DNA cloned from Streptomyces sp. 4353 could also be used as candidate for hybrid and/or heterologous production of AHBA, and the combinatorial biosynthesis of ansamycins.
last changed 2018/11/14 09:26

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