Some molecular properties of asparagine synthetase from rat liver.
Asparagine synthetase purified from rat liver reveals two species (slower migrating band I and faster migrating band II) when subjected to polyacrylamide gel electrophoresis under nondenaturing conditions (S. Hongo and T. Sato (1981) Anal. Biochem. 114, 163-166). We have investigated some molecular properties of these species. Elution of band I from the gel and re-electrophoresis showed that band I yielded band II similar to that of the initial run. Peptide maps by limited proteolysis were very similar and amino acid compositions were also alike in the two species. L-Lysine was identified as the sole NH2-terminal amino acid in both the species. By cross-linking experiments the enzyme was shown to be a dimeric protein. When the purified enzyme was subjected to isoelectric focusing the enzyme activity and protein focused at pH 6.0 in a single peak. These results demonstrate that rat liver asparagine synthetase is composed of two identical subunits. The enzyme, inactivated by storage at -20 degrees C for about 3 months, showed aggregated forms in polyacrylamide gel electrophoresis, and was reactivated markedly by the addition of dithiothreitol.
D000818:Animals; D001217:Aspartate-Ammonia Ligase; D003432:Cross-Linking Reagents; D004229:Dithiothreitol / Q000494:pharmacology; D004789:Enzyme Activation / Q000187:drug effects; D007526:Isoelectric Point; D008025:Ligases; D008099:Liver / Q000201:enzymology*; D046911:Macromolecular Substances; D010446:Peptide Fragments / Q000032:analysis; D051381:Rats
D003432:Cross-Linking Reagents; D046911:Macromolecular Substances; D010446:Peptide Fragments; D008025:Ligases; D001217:Aspartate-Ammonia Ligase; D004229:Dithiothreitol