Differential response of cysteine residues in pig muscle phosphoglucose isomerase to seven sulfhydryl-modifying reagents.
The three cysteine residues per subunit of pig muscle phosphoglucose isomerase show different reactivities toward various sulfhydryl reagents. The organomercurial, p-mercuribenzoate, can titrate two of the sulfhydryl groups under nondenaturing conditions. 2,2'-Dithiodipyridine, 5,5'-dithiobis(2-nitrobenzoic acid), iodoacetamide, methyl 2-pyridyl disulfide, and 2-(2'-pyridylmercapto)mercuri-4-nitrophenol all label only one sulfhydryl group under the same conditions, whereas iodoacetic acid does not react with any of the sulfhydryl groups except when the enzyme is fully denatured. It is concluded, therefore, that charge, rather than steric restraint, is the determining factor for the differences seen in the modification patterns of the enzyme by these reagents. When enzyme was first labeled with 2,2'-dithiodipyridine and subsequently with p-mercuribenzoate, it was found that the latter, in a secondary process, will stoichiometrically react with the anion released by the former after the initial reaction with cysteine. The differences in reactivity of the cysteine residues toward the referred-to reagents have been exploited to specifically modify each of the three individual cysteine residues of pig muscle phosphoglucose isomerase.
D000595:Amino Acid Sequence; D000818:Animals; D003545:Cysteine / Q000032:analysis*; D005956:Glucose-6-Phosphate Isomerase / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D066298:In Vitro Techniques; D009132:Muscles / Q000201:enzymology*; D011489:Protein Denaturation; D013439:Sulfhydryl Reagents / Q000494:pharmacology*; D013552:Swine
D013439:Sulfhydryl Reagents; D005956:Glucose-6-Phosphate Isomerase; D003545:Cysteine