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The proteins encoded by the rbs operon of escherichia coli : I. Overproduction, purification, characterization, and functional analysis of RbsA
Author(s) -
Barroga Charlene F.,
Zhang Huide,
Wajih Nadeem,
Bouyer James H.,
Hermodson Mark A.
Publication year - 1996
Publication title -
protein science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 3.353
H-Index - 175
eISSN - 1469-896X
pISSN - 0961-8368
DOI - 10.1002/pro.5560050611
Subject(s) - escherichia coli , operon , overproduction , biochemistry , biology , function (biology) , recombinant dna , lac operon , gtp' , atpase , gene , nucleotide , enzyme , microbiology and biotechnology , chemistry , genetics
The nucleotide‐binding component of the high‐affinity ribose transport system of Escherichia coli , RbsA, was overproduced from a T7–7 expression vector, and the protein was purified. Biochemical analyses of the purified protein indicated that the ATP analogues, 5′‐FSBA and 8‐azido ATP, covalently labeled the protein, a reaction that was inhibited by ATP, but not by GTP or CTP. The pure protein exhibited low‐level ATPase activity with a K m of about 140 μM. Analyses of bacterial strains carrying chromosomal deletions of rbsA and other rbs genes suggested that RbsA is important for the chemotaxis function, a surprising result that was not anticipated from previous studies. However, an inconsistency between the several results from deletion strains raises questions regarding the interpretations of the in vivo data.