Subunit interactions in aspartate transcarbamylase. Characterization of a complex between the catalytic and the regulatory subunits.
The complex formed when excess regulatory subunits (r2) of aspartate transcarbamylase is added to a dilute solution of the catalytic subunit (c3) has been further studied. By stabilizing the complex with saturating levels or r2, it was possible to perform ultracentrifugation in sucrose density gradients. The sedimentation coefficient of the complex (7.7 plus or minus 0.2 S) is intermediate between those of the catalytic subunit (5.8 S) and of the native enzyme (11.7 S). Consideration of the likely hydrodynamic properties of the complex suggests that this sedimentation coefficient may be consistent with the c3r6 structure previously proposed. The formation of c3r6 from c3 and r2 is readily reversible. At nonsaturating levels or r2, conversion to the native enzyme (c3r6) takes place. This conversion is inhibited by high concentrations of r2. The c3r6 complex shows Michaelis-Menten kinetics with a low Km for aspartate and considerable substrate inhibition. The pH activity profile at high aspartate concentrations is almost identical with that of the native enzyme. All of these observations suggest that c3r6 represents the relaxed (R) state of aspartate transcarbamylase. The insensitivity of c3r6 toward CTP or ATP can also be explained by considering c3r6 as a stabilized relaxed state. These properties of c3r6 have significant implications regarding the allosteric mechanism of the native enzyme.
D000255:Adenosine Triphosphate / Q000494:pharmacology; D000494:Allosteric Regulation; D001221:Aspartate Carbamoyltransferase / Q000378:metabolism; D001665:Binding Sites; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D003597:Cytosine Nucleotides / Q000494:pharmacology; D004926:Escherichia coli / Q000201:enzymology; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D046911:Macromolecular Substances; D011485:Protein Binding; D011487:Protein Conformation
D003597:Cytosine Nucleotides; D046911:Macromolecular Substances; D000255:Adenosine Triphosphate; D001221:Aspartate Carbamoyltransferase