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1.
Int J Biol Macromol ; 18(4): 255-62, 1996 Jun.
Article in English | MEDLINE | ID: mdl-8739129

ABSTRACT

Hypoxanthine-guanine phosphoribosyltransferase (HGPRT, E.C.2.4.2.8) from Artemia cysts exhibits maximum activity at 70 degrees C. Its thermal stability has been examined following enzymatic activity as a function of temperature. Cold-induced renaturation experiments of samples heated at increasing temperatures showed that reversibility of thermal inactivation depends on the incubation time and final temperature. Prolonged incubation of the thermoinactivated enzyme at 0 degree C did not afford any further increase of the catalytic activity at 37 degrees C. The complex substrate PRPP:Mg protects HGPRT from thermal inactivation. However, incubations with hypoxanthine rendered a less thermostable enzyme at any temperature tested. The irreversible inactivation of HGPRT proceeds in two exponential steps. The analysis of the apparent rate constants for the fast and the slow phases, lambda 1 and lambda 2 as per the Lumry and Eyring model suggests the existence of more than three states in the thermal denaturation pathway of the free enzyme. In the presence of PRPP:Mg the irreversible process follows a single exponential and proceeds very slowly below 70 degrees C. PRPP:Mg also protects the enzyme from inactivation by NEM and pCMB, suggesting that -SH groups may be in the vicinity of the active site.


Subject(s)
Artemia/enzymology , Hypoxanthine Phosphoribosyltransferase/antagonists & inhibitors , Hypoxanthine Phosphoribosyltransferase/metabolism , Animals , Binding Sites , Enzyme Stability , Hypoxanthine Phosphoribosyltransferase/chemistry , Kinetics , Macromolecular Substances , Phosphoribosyl Pyrophosphate , Protein Denaturation , Protein Folding , Substrate Specificity , Temperature
2.
Rev Esp Cardiol ; 49(4): 281-7, 1996 Apr.
Article in Spanish | MEDLINE | ID: mdl-8650405

ABSTRACT

OBJECTIVE: This study was designed to investigate urate production by swine hearts using an in vivo regionally ischemic-reperfused model. ANIMALS AND METHODS: Ten female pigs underwent 60 minutes of myocardial ischemia by clamping of the left anterior descending artery and afterwards 120 minutes of reperfusion. Epicardial biopsies and blood samples from coronary sinus were taken before ligation, at the end of ischemic period and 5, 30, 60 and 120 minutes upon reperfusion. RESULTS: During ischemia, tissue levels of ATP and ADP greatly declined with a subsequent increase in the concentration of AMP, inosine and hypoxanthine (33 +/- 12 vs 93 +/- 17, 26 +/- 8 vs 768 +/- 86 and 32 +/- 10 vs 219 +/- 26 nmol/g dry weight, p < 0.01 for each). Despite the great increase in the hypoxanthine levels, uric acid concentration remained constant (69 +/- 9 vs 32 +/- 12 nmol/g dry weight, NS). Hypoxanthine, xanthine and uric acid concentrations increased in blood samples obtained from the coronary sinus at the end of ischemic period (17.99 vs 31.03 nmol/ml, p < 0.01, 0.29 vs 1.45 nmol/ml, p < 0.05 and 1.20 vs 2.31 nmol/ml, p < 0.01 respectively) and were enhanced upon reperfusion (35.8 and 3.89 nmol/ml for hypoxanthine and uric acid respectively, p < 0.05) without any significant modifications in their concentrations at the arterial level. CONCLUSION: These results demonstrate that the ischemic-reperfused swine heart produces urate probably outside the myocardium.


Subject(s)
Myocardial Reperfusion Injury/metabolism , Uric Acid/metabolism , Animals , Female , Free Radicals , Hypoxanthine , Hypoxanthines/blood , Myocardial Reperfusion Injury/blood , Myocardium/metabolism , Swine , Uric Acid/blood , Xanthine , Xanthines/blood
3.
Biochim Biophys Acta ; 1033(1): 114-7, 1990 Jan 29.
Article in English | MEDLINE | ID: mdl-1689183

ABSTRACT

5'-Phosphoribosylpyrophosphate amidotransferase, which catalyzes the synthesis of phosphoribosylamine in the de novo synthesis of purine nucleotides, has been detected and partially purified approx. 800-fold from Artemia sp. nauplii. The apparent Km values for 5'-phosphoribosyl 1-pyrophosphate as substrate were 0.7 mM and 0.4 mM in the presence of glutamine and ammonia as nitrogenous sources, respectively, and the enzymatic activity was inhibited by purine 5'-ribonucleotide compounds and 5', 5'''-p1, p4-diguanosine tetraphosphate.


Subject(s)
Amidophosphoribosyltransferase/metabolism , Artemia/enzymology , Pentosyltransferases/metabolism , Amidophosphoribosyltransferase/antagonists & inhibitors , Amidophosphoribosyltransferase/isolation & purification , Ammonia/pharmacology , Animals , Artemia/growth & development , Chromatography , Glutamine/pharmacology , Kinetics , Phosphoribosyl Pyrophosphate/metabolism , Purine Nucleotides/pharmacology
4.
FEBS Lett ; 255(2): 358-60, 1989 Sep 25.
Article in English | MEDLINE | ID: mdl-2477280

ABSTRACT

De novo synthesis of total and ribosomal ribonucleic acids has been studied during the early stages of Artemia sp. development. By in vivo incorporation studies of [14C]HCO3- an increase has been found in both total and ribosomal RNA synthesis post hatching, with a similar distribution of radioactivity and base composition.


Subject(s)
Artemia/growth & development , RNA, Ribosomal/biosynthesis , RNA/biosynthesis , Animals , Base Composition , Bicarbonates/metabolism , RNA/isolation & purification , RNA, Ribosomal/isolation & purification
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