Effect of pH and KCl on aggregation state and sulphydryl groups reactivity of rat skeletal muscle AMP deaminase.
The effects of pH and KCl on sedimentation properties and SH groups reactivity of rat skeletal muscle AMP deaminase have been investigated. The values obtained for apparent molecular weight are consistent with an association of AMP deaminase subunits in response to increasing KCl concentration. Increasing pH value from 6.0 to 8.0 causes a reduction in the apparent molecular weight of the enzyme at high KCl concentration, which can be interpreted as due to a deprotonation-induced isomerization process. Removal of Zn2+ from AMP deaminase has effect similar to alkalinization in modifying the sedimentation properties of the enzyme. In the native enzyme at high K+ concentration about 7, 9 and 12 SH groups can be titrated with Nbs2, approximately 1, 2 and 4 SH groups reacting as fast sets, at pH 6.0, 7.0 and 8.0, respectively. Substitution of the 12 SH groups reactive with Nbs2 at pH 8.0 has no effect on the pH-dependent allosteric behaviour of the enzyme. Removal of K+ causes considerable changes in the reactivity of AMP deaminase towards Nbs2, unmasking a class of additional SH groups, so that the total number of titratable SH groups approaches that of 30 determined in denaturing conditions. In the enzyme previously treated with N-ethylmaleimide to alkylate the fast reacting class of SH groups, the class of additional SH groups are substituted by Nbs2 at basic pH, but not at acidic pH, with a concomitant reduction of the enzyme activity.
D000659:AMP Deaminase / Q000378:metabolism*; D000818:Animals; D002499:Centrifugation, Density Gradient; D004492:Edetic Acid / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008970:Molecular Weight; D009132:Muscles / Q000201:enzymology*; D009709:Nucleotide Deaminases / Q000378:metabolism*; D011189:Potassium Chloride / Q000494:pharmacology*; D011487:Protein Conformation; D051381:Rats; D013438:Sulfhydryl Compounds / Q000378:metabolism; D015032:Zinc / Q000378:metabolism
D013438:Sulfhydryl Compounds; D011189:Potassium Chloride; D004492:Edetic Acid; D009709:Nucleotide Deaminases; D000659:AMP Deaminase; D015032:Zinc