CLONING AND SEQUENCING OF A GENE ENCODING A GLUTAMATE AND ASPARTATE CARRIER OF ESCHERICHIA-COLI K-12

被引:47
作者
WALLACE, B [1 ]
YANG, YJ [1 ]
HONG, JS [1 ]
LUM, D [1 ]
机构
[1] BOSTON BIOMED RES INST,BOSTON,MA 02114
关键词
D O I
10.1128/jb.172.6.3214-3220.1990
中图分类号
Q93 [微生物学];
学科分类号
071005 ; 100705 ;
摘要
A gene encoding a carrier protein for glutamate and aspartate was cloned into Escherichia coli K-12 strain BK9MDG by using the high-copy-number plasmid pBR322. The gene (designated gltP) is probably identical to a gene recently cloned from E. coli B (Y. Deguchi, I. Yamato, and Y. Anraku, J. Bacteriol. 171:1314-1319). A 1.6-kilobase DNA fragment containing gltP was subcloned into the expression plasmids pT7-5 and pT7-6, and its product was identified by a phage T7 RNA polymerase-T7 promoter coupled system (S. Tabor and C. C. Richardson, Proc. Natl. Acad. Sci. USA 82:1074-1078) as a polypeptide with an apparent mass of 38 kilodaltons. A portion of the gltP polypeptide was associated with the cytoplasmic membrane. The nucleotide sequence of the 1.6-kilobase fragment was determined. It contained an open reading frame capable of encoding a highly hydrophobic polypeptide of 395 amino acids, containing four possible transmembrane segments. Uptake of glutamate and aspartate was increased 5.5- and 4.5-fold, respectively, in strains containing gltP plasmids. Glutamate uptake was insensitive to the concentration of Na+ and was inhibited by L-cysteate and β-hydroxyaspartate. These results suggest that gltP is a structural gene for a carrier protein of the Na+-independent, binding-protein-independent glutamate-aspartate transport system.
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页码:3214 / 3220
页数:7
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