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鼠伤寒沙门氏菌cys E和cys M基因在哺乳动物细胞中的共表达:通过转基因在绵羊中建立半胱氨酸生物合成的一步。

Coexpression of the cys E and cys M genes of Salmonella typhimurium in mammalian cells: a step towards establishing cysteine biosynthesis in sheep by transgenesis.

作者信息

Sivaprasad A V, Kuczek E S, Bawden C S, Rogers G E

机构信息

Biochemistry Department, University of Adelaide, Australia.

出版信息

Transgenic Res. 1992 Mar;1(2):79-92. doi: 10.1007/BF02513025.

Abstract

The Salmonella typhimurium genes for serine acetyltransferase (cys E) and O-acetylserine sulphydrylase B (cys M) were isolated and characterized in order to express these as transgenes in sheep to establish a cysteine biosynthesis pathway and, thereby, to achieve an increased rate of wool growth. Comparison of the S. typhimurium and Escherichia coli genes showed considerable homology, both at the nucleotide and amino acid sequence levels. The in vitro and in vivo expression studies showed that both genes could be transcribed and translated in eukaryotic cells and that their products could function as active enzymes. The cys M gene of S. typhimurium possessed a GUG initiation codon, like its E. coli counterpart, but translation could be initiated using this codon in eukaryotic cells to give an active enzyme product. Chinese hamster ovary cells, stably transfected with a tandem arrangement of the two genes, showed a capacity to synthesize cysteine in vivo, indicating the establishment of a cysteine biosynthesis pathway in these cells. The measured levels of activity of the gene products suggest that improved wool growth is possible by transgenesis of sheep with these genes.

摘要

鼠伤寒沙门氏菌中丝氨酸乙酰转移酶(cys E)和O - 乙酰丝氨酸巯基酶B(cys M)的基因被分离并进行了表征,目的是将这些基因作为转基因在绵羊中表达,以建立半胱氨酸生物合成途径,从而提高羊毛生长速率。鼠伤寒沙门氏菌和大肠杆菌基因的比较表明,在核苷酸和氨基酸序列水平上都有相当高的同源性。体外和体内表达研究表明,这两个基因都可以在真核细胞中转录和翻译,并且它们的产物可以作为活性酶发挥作用。鼠伤寒沙门氏菌的cys M基因与大肠杆菌的对应基因一样,具有一个GUG起始密码子,但在真核细胞中使用这个密码子可以起始翻译,产生有活性的酶产物。用这两个基因的串联排列稳定转染的中国仓鼠卵巢细胞显示出在体内合成半胱氨酸的能力,表明在这些细胞中建立了半胱氨酸生物合成途径。所测基因产物的活性水平表明,用这些基因对绵羊进行转基因有可能改善羊毛生长。

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