Rat muscle 5' adenylic acid aminohydrolase. I. Purification and subunit structure

AMP deaminase (AMP aminohydrolase, EC 3.5.4.6) has been purified to apparent homogeneity from rat muscle. The preparation exhibits a single polypeptide band with a molecular weight of 60,000 on polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate. The enzyme has a sedimentation coefficent of 11.3 S. Analysis by sedimentation equilibrium techniques showed the native enzyme to have a molecular weight of 238,000, whereas the enzyme, when analyzed in 6 M guanidine hydrochloride and 10 mM 2 mercaptoethanol, had a molecular weight of only 59,500. The amino acid composition of the enzyme was determined and peptide mapping was performed on a tryptic digest of S carboxymethylated enzyme. NH₂ terminal analysis by both the dansylation and cyanate procedures failed to identify a free NH₂ terminus. Treatment of the enzyme with carboxypeptidase A resulted in the release of approximately 0.5 mol each of valine and leucine per 60,000 g of enzyme. The data presented indicate that the native enzyme has a tetrameric structure consisting of four polypeptide chains each having a molecular weight of 60,000. The COOH terminal analysis can be interpreted either as an indication of subunit heterogeneity or as a result of incomplete digestion of a X Leu Val sequence at the end of a single type of polypeptide chain. Tryptic peptide maps strongly support the latter interpretation and suggest that the subunits are essentially identical.