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Title: Biosynthesis of spectinomycin from Streptomyces spectabilis and heterologous expression
Keywords: S. venezuelae YJ003. S. kanamyceticus, S. spectabilis, Myo-inositol synthase, Myo-inositol dehydrogenase, aminotransferase, Heterologous hosts, Streptamine, Geneticall engineered aminoglycosides antibiotics, N-methlated kanamycin.
Issue Date: 27-Mar-2019
Abstract: Spectinomycin is an aminocyclitol antibiotic produced by Streptomyces spectabils and may be regarded as part of a subclass of aminoglycoside antibiotics. The spectinomycin is composed of spectinamine which is derived from myo-inositol and actinospectose derived from TDP-glucose. It is bacteriostatic and exerts its antibacterial effect by binding to the 30S ribosome which disrupts bacterial protein synthesis. Spectinomycin is primarily effective against Gram-negative bacteria such as Pasteurella haemolytica, Pasteurella multocida and Haemophilus somnus. It has limited activity against a number of Grampositive bacteria. Generally, it is not active against anaerobic bacteria. The analysis of 17 kb nucleotide sequence of spectinomycin biosynthesis gene cluster of S. spectabilis revealed 16 open reading frames (ORFs) and which include biosynthetic genes, resistance genes and regulatory genes. Among them, three sepctinomycin biosynthesis genes myo-inositol synthase (spcA), myo-inositol dehydrogenase (spcB) and aminotransferase (spcS2) were demonstrated by in vivo assay in S. venezuelae YJ003 and are sufficient for the biosynthesis of streptamine because myo-inositol dehydrogenase and aminotransferase has dual character. The production of the sreptamine was confirmed by ESI-mass, LC-mass and HPLC profile, respectively. Similarly, the production of genetically engineered N-methylated kanamycin by in vivo assay of gene encoding a protein that catalyzes the N-methylation of streptamine in biosynthetic pathway of spm in Streptomyces kanamyceticus. The production of the methylated kanamycin was checked by ESI-mass, tandem Mass-Mass spectrometry, HPLC and MIC test, respectively. The heterologous expression of the SPC8 cosmid, contining the spectinomycin biosynthetic gene cluster was carried out in S. venezuelae YJ003 and the production of spectinomycin was confirmed by biological assay, ESI-mass spectrometry, LC-mass and high pressure liquid chromatography (HPLC) of isolated secondary metabolites from S.
Appears in Collections:500 Natural sciences and mathematics

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