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基因工程丙酰螺旋霉素 预览 被引量:5

A GENETICALLY ENGINEERED HYBRID ANTIBIOTIC PROPIONYLSPIRAMYCIN
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摘要 以碳霉素4"异戊酰基转移酶基因(CarE)为探针,从麦迪霉素产生菌基因文库中,经菌落杂交获得阳性菌落pCN10F5。经分子杂交证明pCN10F5DNA的BamHI-BamHI8.0kb片段与CarE基因同源。将pCN10F5 BamHI8.0kb片段分别克隆到pIJ680、pWHM3,pWHM601质粒载体上,获得重组质粒p6F5,pWF5和PGF5。含上述重组质粒的螺旋霉素产生菌克隆菌株均产生与 A genetically engineered hybrid antibioticPropionylspiram-ycin producing strain was constructed by expression of Midecamycin 4″- propionyltransferase(MPT)gene in the Spiramycin producing strain (S. sp- iramyceticus),The MPT gene was identified from the genomic library ofMidecamycin producing S,mycarofaciens 1748 strain using S. ihermotole-rans 4″-isovaleryltransferase(CarE)gene as a probe,Southern blot hybri-dization of the pCN10F5 cosmid clone localized the CarE homology in an8.0kb BamHI DNA fragment.This fragment was subcloned into plasmidspIJ680,pWHM3 and pWNM601 to give respective recombinant plasmidsp6F5,pWF5 and pGF5.Transfomation of S. spiramyceticus and S. livid- ans with these plasmids resulted in production or conversion of exogenou-sly added Spiramycin to a hybrid antibiotic,Based on their physicochem-ical properties and spectral evidences the main components were 4″propio-nylspiramycin Ⅱ&Ⅲ. Southern hybridization confirmed that the BamHI-BamHI 8.0kb fragment was cloned in the the Spiramycin producing stra-in. Restriction map of BamHI8.0kb fragment was constructed.The MPTgene was localized on a EcoRI-EcoRI-pstI 2.65kb fragment through Sou-thern hybridization, subcloning and its expression in S. lividans as wellas in S. spiramyceficus.The nucleotide sequeiice of the MPT gene hasbeen determined.The codon utilization program predicts that the codingregion for MPT contains 1164 nucleotide,which encodes a protein of 388amino acids,with ATG as a start codon and TGA as a terminator cod-on,The deduced amino acids sequence of MPT shows 67.9% identity wi-th CarE amino acids sequence,The genetically engineered Propionylspira-mycin producing strain was very stable when put into successive passageson slant medium or in batchwise fermentation.
作者 王以光 金莲舫 Wang Yi-guang;Jin Lian-feng;Xu Xiao-min;Chang Xu-lun and Zeng Yin(Institute of Medicinal Biotechnology,Chinese Academy of Medical Sciences and Peking Union Medical College,Beijng 100050)
出处 《中国抗生素杂志》 CAS CSCD 北大核心 1994年第2期 109-116,共8页 Chinese Journal of Antibiotics
关键词 基因工程 抗生素 丙酰螺旋霉素 Midccamycin 4″hydroxyl propionyltransferase gene Genetically engineered hybrid antibioticsi 4″propionylspiramycin
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