Site-specific alteration of cysteine 176 and cysteine 234 in the lactose carrier of Escherichia coli.
In the present study, Cys-176 and Cys-234 in the lactose carrier have been modified to serine residues via site-specific mutagenesis. The resultant mutants have been characterized with regard to galactoside transport activity and sulfhydryl reagent sensitivity. The mutant proteins (in which Cys-176 or Cys-234 had been replaced with serine) are able to effectively transport galactosides, although the transport rates for lactose and methyl-beta-D-galactopyranoside are slightly reduced compared to the normal lactose carrier. In addition, both mutants are less sensitive than the wild-type to high concentrations of two different sulfhydryl reagents, N-ethylmaleimide and p-hydroxymercuribenzoate. Overall, the data are consistent with the idea that Cys-176 and Cys-234 are close to the substrate recognition site. However, neither residue appears to be essential for galactoside transport by providing an ionizable group near the active site or by forming a disulfide bond.
D003545:Cysteine; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D029968:Escherichia coli Proteins; D005033:Ethylmaleimide / Q000494:pharmacology; D006902:Hydroxymercuribenzoates / Q000494:pharmacology; D007700:Kinetics; D007785:Lactose / Q000378:metabolism; D026901:Membrane Transport Proteins / Q000235:genetics* / Q000378:metabolism; D009004:Monosaccharide Transport Proteins; D009154:Mutation; D027981:Symporters
D029968:Escherichia coli Proteins; D006902:Hydroxymercuribenzoates; C050221:LacY protein, E coli; D026901:Membrane Transport Proteins; D009004:Monosaccharide Transport Proteins; D027981:Symporters; C030601:4-hydroxymercuribenzoate; C021499:lactose permease; D007785:Lactose; D003545:Cysteine; D005033:Ethylmaleimide