Activation of rabbit cardiac AMP aminohydrolase by ADP: A component of a mechanism guarding against ATP depletion

The kinetic properties of a 300-fold purified cardiac AMP deaminase were studied and compared with those of the corresponding enzyme from skeletal muscle. The heart enzyme is activated by ATP and less efficiently by ADP, and is inhibited by Pi, phosphocreatine and GTP. ATP, even at micromolar concentrations, is able to abolish the effects of the inhibitors. The affinity of the enzyme for AMP is low in the absence of activators (Km 3.1 mM), but, in the presence of ATP, becomes as high as that of skeletal-muscle AMP deaminase (Km 0.4 mM). The maximal activation by ATP is observed at alkaline pH (pH 7.5-8.0). Under the same conditions ATP is maximally inhibitory for skeletal-muscle enzyme. These results suggest that AMP deaminase in the heart is always in the activated state, whereas in skeletal muscle the enzyme is active only during exhaustive contractions.

Section: Effect Of Ph and Mg2+ Ions On The Activity And Speculations supporting

confidence: 82%

Adenylate metabolism in the heart. Regulatory properties of rabbit cardiac adenylate deaminase

Barsacchi¹,

Ranieri-Raggi²,

Bergamini³

et al. 1979

“…This is proposed to occur during periods of net ATP hydrolysis, by pulling the myokinase reaction (2 ADP-ATP+AMP) toward ATP production [9][10][11][12]. The activation of AMP deaminase by decreasing energy charge [10,12] and the initiation of AMP deamination in the highly aerobic fast-twitch red fibres at very high work loads [13] support this proposal.…”

Section: Introductionsupporting

confidence: 52%

Release of purine and pyrimidine nucleosides and their catabolites from the perfused rat hindlimb in response to noradrenaline, vasopressin, angiotensin II and sciatic-nerve stimulation

Clark

Richards

Hettiarachchi

et al. 1990

Uric acid and uracil were released at constant rates (0.95 and 0.4 nmol/min per g respectively) by the perfused rat hindlimb. Noradrenaline, vasopressin or angiotensin II further increased the release of these substances 2-5-fold, coinciding with increases in both perfusion pressure (vasoconstriction) and 02 uptake. The hindlimb also released, but in lesser amounts, uridine, hypoxanthine, xanthine, inosine and guanosine, and all but hypoxanthine and guanosine were increased during intense vasoconstriction. Uric acid and uracil releases were increased by noradrenaline in a dose-dependent manner. However, the release of these substances did not fully correspond with the dose-dependent increase in 02 uptake and perfusion pressure, where changes in the latter occurred at lower doses of noradrenaline. Sciatic-nerve stimulation (skeletalmuscle contraction) did not increase the release of uracil, uric acid or uridine, but instead increased the release of inosine (7-fold) and hypoxanthine (2-fold). Since the UTP content as well as the UTP/ATP ratio are higher in smooth muscle than in skeletal niuscle, it is proposed that release of uric acid and uracil arises from increased metabolism of the respective adenosine and uridine nucleotides during intense constriction of smooth muscle.

“…For example, we demonstrate that the Km of purified rabbit heart AMP deaminase was decreased 3.4-fold by 1.0 mM ATP, whereas a 7-fold decrease was reported for a crude preparation of this enzyme [29]. Non-homogeneous cardiac AMP deaminase exhibited greater sensitivity to millimolar ADP than ATP [28], whereas the isolated rabbit heart enzyme was somewhat more sensitive to ATP (Figure 4). At least some of these discrepancies may reflect contamination ofcrude AMP deaminase preparations by nucleoside-diphosphate kinase (ATP: nucleoside-diphosphate phosphotransferase, EC 2.7.4.6) and/or adenylate kinase (ATP: AMP phosphotransferase, EC 2.7.4.3.…”

Section: Enriched Cardiac Amp Deaminase Preparations Have Reportedmentioning

confidence: 76%

“…The activity of isolated rabbit heart AMP deaminase was modulated by several effectors: ADP, ATP, GTP, inorganic (poly)phosphate compounds and fatty acyl-CoAs. Allosteric activation by ADP and ATP and inhibition by GTP, phosphates and fatty acyl-CoA have been observed in enriched cardiac AMP deaminase preparations [23,24,27,28] and with the isolated skeletal-muscle enzyme [37,38]. Quantitatively, however, variable sensitivities toward each effector have been reported.…”

Section: Enriched Cardiac Amp Deaminase Preparations Have Reportedmentioning

confidence: 99%

Isolation and characterization of AMP deaminase from mammalian (rabbit) myocardium

Thakkar¹,

Janero²,

Yarwood³

et al. 1993

AMP deaminase (AMP aminohydrolase, EC 3.5.4.6) is a ubiquitous enzyme in eukaryotes, which may play a role in ATP catabolism during myocardial ischaemia. We report isolation of AMP deaminase from rabbit myocardium with a 19% recovery and a 650-fold enrichment, using a newly devised protocol involving sequential cation-exchange, gel-permeation and affinity chromatographies. The cardiac AMP deaminase preparation described was electrophoretically and chromatographically homogeneous and contained one unique N-terminal residue (leucine). The isolated enzyme was sensitive to various cations (K+, Mg2+, Ca2+). The pH optimum of purified cardiac AMP deaminase was 6.8, its pI was 6.5, and it displayed substrate-specificity toward 5'-AMP. The subunit molecular mass of rabbit heart AMP deaminase on SDS/PAGE (81 kDa) and the holoenzyme molecular mass as estimated by non-denaturing size-exclusion h.p.l.c. (330 kDa) indicated that the native enzyme was a tetramer. Cardiac AMP deaminase displayed a sigmoidal substrate-saturation curve in the presence of 100 mM KCl. Apparent Michaelis constants were a Km of 5.8 mM AMP and a Vmax. of 11.1 mumol/min per mg of protein. ATP and ADP were positive allosteric effectors of cardiac AMP deaminase: the apparent Km was decreased to 1.7 mM by 1.0 mM ATP. The enzyme was inhibited by GTP, coformycin, coformycin 5'-phosphate, palmitoyl-CoA, inorganic phosphate compounds, and the metal chelator o-phenanthroline. No inhibition either by product nucleotide (IMP) or by nicotinamide nucleotides was detected when these agents were examined at concentrations up to 2.5 mM. We conclude that this enzyme preparation offers a means by which the kinetic mechanism and regulation of mammalian cardiac AMP deaminase may be directly investigated.