AMP deaminase from yeast. Role in AMP degradation, large scale purification, and properties of the native and proteolyzed enzyme.

摘要: Abstract Eukaryotes have been proposed to depend on AMP deaminase as a primary step in the regulation of intracellular adenine nucleotide pools. This report describes 1) role adenylate metabolism yeast cell extracts, 2) method for large scale purification enzyme, 3) kinetic properties native and proteolyzed enzymes, 4) reaction mechanism, 5) regulatory interactions with ATP, GTP, MgATP, ADP, PO4. Allosteric is physiological significance, since expression gene constitutive (Meyer, S. L., Kvalnes-Krick, K. Schramm, V. L. (1989) Biochemistry 28, 8734-8743). The ATP cell-free extracts demonstrates that sole pathway catabolism these extracts. Purification enzyme from bakers' yields proteolytically cleaved Mr 86,000, which missing 192 amino acids N-terminal region. Extracts Escherichia coli containing plasmid contained only unproteolyzed 100,000. highly unstable during purification. Substrate saturation plots are sigmoidal. In presence allosteric activator, exhibits normal kinetics. activates by increasing affinity 1.3 0.2 mM without affecting VM. Activation more efficient than half-maximum activation constants 6 80 microM, respectively. similar. Thus, region not required catalysis or activation. competitively inhibited GTP PO4 respect AMP. inhibition inhibitors decrease ATP. therefore, tightens binding PO4, products reaction, NH3 IMP, competitive against substrate, consistent rapid equilibrium random mechanism. Kinetic dissociation reported binary ternary substrate product complexes modulators.