Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.1.3.5 (5'-nucleotidase)
 3,167 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have studied the effects of phorbol 12-myristate 13-acetate (PMA, 15 micrograms) on pulmonary endothelial ectoenzyme [angiotensin-converting enzyme (ACE) and 5'-nucleotidase (NCT)] function in isolated rabbit lungs perfused in situ with platelet-poor (PPP) or platelet-rich (PRP) plasma in the presence or absence of neutrophils. Enzyme activities were estimated from the hydrolysis of the substrates [3H]benzoyl-Phe-Ala-Pro ([3H]BPAP) by ACE and 14C-labeled AMP by NCT during a single transpulmonary passage, using indicator-dilution techniques. In all treatment groups PMA produced a delayed increase in pulmonary vascular resistance to about three times the control value. PMA alone [in lungs perfused with PPP (n = 5 animals) or PRP (n = 6)] or neutrophils alone (in PPP-perfused lungs, n = 5) had no effect on enzyme activity. However, PMA-activated neutrophils (n = 5) decreased percent metabolism (%M) of [3H]BPAP from 87 +/- 3 to 77 +/- 4% (30 min after PMA), and the apparent first-order parameter [ratio of maximum activity to Michaelis constant (Amax/Km)] for ACE from 821 +/- 114 to 613 +/- 61 ml/min (30 min after PMA). At the same time, Km values of BPAP for ACE and AMP for NCT were elevated from 9.2 +/- 2.2 to 19.3 +/- 3 microM and 6.7 +/- 1.2 to 15.1 +/- 3.6 microM, respectively, whereas Amax (product of enzyme mass and rate of product formation, thus an index of perfused microvascular surface area) did not change.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:PMA-activated neutrophils decrease pulmonary endothelial ectoenzyme activities in perfused rabbit lungs. 133 99

We investigated the early phase of pulmonary endothelial injury in rabbits exposed to a single dose (30 Gy) of ionizing radiation to the chest, by measuring endothelium-bound ectoenzyme activities. Utilizing multiple indicator-dilution techniques, the metabolism of [3H]benzoyl-Phe-Ala-Pro (BPAP) and [14C]5'-AMP by angiotensin-converting enzyme (ACE) and 5'-nucleotidase (NCT), respectively, was studied during a single transpulmonary passage in conscious, chronically catheterized rabbits. From these data, the apparent kinetic constants Km and Amax were calculated. A significant (p less than 0.05) decrease in the metabolism of trace amounts of BPAP and 5'-AMP was observed at 2, 24, and 48 hr after irradiation. A similar decrease in the apparent first order rate constant (Amax/Km) of ACE was observed at 2 hr, but returned to control levels by 24 and 48 hr after irradiation. Apparent Km values of ACE for BPAP and NCT for 5'-AMP were elevated at 2, 24, and 48 hr post-treatment, whereas Amax (product of enzyme mass and the constant of product formation, kcat) of ACE was elevated at 2 and 24 hr but not at 48 hr, and Amax for NCT was elevated at 2 hr post-treatment only. Significant decreases in mean arterial blood pressure and pulmonary blood flow (Qb) at 2 hr post-treatment, and increases in Qb at 24 and 48 hr post-treatment were also recorded. No changes in endothelial structure were observed 2 hr after irradiation at the light or electron microscope level. We conclude that the early phase of radiation-induced lung injury includes changes in endothelial enzyme function in the absence of structural damage, as reflected in an apparent decrease in affinity of ACE and NCT for their substrates, allowing for the possibility that hemodynamic disturbances or their sequalae could also have contributed to the decrease in enzyme function.
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PMID:Early effects of ionizing radiation on pulmonary endothelial angiotensin-converting enzyme and 5'-nucleotidase, in vivo. 284 Jul 53

We investigated changes in angiotensin converting-enzyme (ACE) activity before and at 5, 15, 60, and 240 min after 20 micrograms phorbol myristate acetate/kg body wt iv in conscious rabbits. ACE activity was estimated in vivo from the single-pass transpulmonary metabolism of the synthetic substrate [3H]benzoyl-Phe-Ala-Pro [( 3H]BPAP) under first-order reaction conditions. Within 5 min after PMA administration, all animals developed profound granulocytopenia (15% of control) and moderate thrombocytopenia (57% of control), both lasting for the duration of the experiment. Concomitantly, there was a significant decrease in the transpulmonary metabolism of [3H]BPAP and the calculated apparent first-order reaction constant Amax/Km of ACE for [3H]BPAP. No histological evidence of lung injury was observed at these times. Since a concomitant fall in the permeability surface area product for urea was also observed, we considered that the apparent decline in ACE activity might have resulted from a reduction in perfused endothelial surface area. To resolve this, we studied the effect of PMA on the Km (a measure of enzyme affinity for its substrate) and Amax (a derivative of Vmax that is dependent upon total enzyme present and thus capillary surface area) of ACE and 5'-nucleotidase for [3H]BPAP and [14C]AMP, respectively. A significant increase in Km for both enzymes was observed at 1 h after PMA, whereas Amax was unaffected, suggesting that low-dose PMA may indeed produce endothelial cell enzyme dysfunction independent of its effect on capillary surface area. These results provide evidence of pulmonary capillary functional injury before or in the absence of structural endothelial damage.
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PMID:Early pulmonary endothelial enzyme dysfunction after phorbol ester in conscious rabbits. 369 30

Angiotensin-converting enzyme and 5'-nucleotidase line the luminal surface of pulmonary microvascular endothelium and participate in the synthesis and/or degradation of potent vasoactive substances. We applied Michaelis-Menten kinetics in simultaneous estimations of apparent constants Km and Amax (product of Vmax and microvascular plasma volume) of these two enzymes for the substrates 3H-labeled benzoyl-Phe-Ala-Pro and 14C-labeled 5'-AMP, respectively, in vivo. Values of angiotensin-converting enzyme for benzoyl-Phe-Ala-Pro (Km = 10-11 microM; Amax = 12-13 mumol X min-1) were somewhat higher than published estimates in vitro and changed predictably in response to the known enzyme inhibitor captopril. Kinetic values of 5'-nucleotidase for 5'-AMP (Km = 3-4 microM; Amax = 3-4 mumol/min) were substantially lower than those reported in vitro but also responded predictably to the competitive inhibitor of 5'-nucleotidase, adenosine 5'-[alpha, beta-methylene]diphosphate. These data offer in vivo estimates of enzyme kinetics that are useful in revealing enzyme behavior in their normal physiological environment and provide means of evaluating the action of pharmacological, physiological, and pathological modulators of enzyme activity, in vivo.
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PMID:Kinetics of pulmonary angiotensin-converting enzyme and 5'-nucleotidase in vivo. 609 4

We monitored the activity of pulmonary microvascular endothelial-bound angiotensin-converting enzyme (ACE) in vivo by means of multiple indicator-dilution-type techniques, utilizing three different probes: the hydrolysis of two substrates, [3H]-benzoyl-Phe-Ala-Pro (BPAP) and [14C]benzoyl-Ala-Gly-Pro (BAGP), and the binding of the inhibitor [3H]RAC-X-65 (RAC), all measured during a single transpulmonary passage in anesthetized rabbits, placed on total heart bypass, so that both systemic and pulmonary circulations were fully supported by means of a two-channel extracorporeal pump. Experiments were performed at pulmonary blood flows (Qb) of 250, 400, 560, and 800 ml/min in control or indomethacin-pretreated rabbits. ACE activity was also compared to that of pulmonary microvascular endothelial-bound 5'-nucleotidase, by measuring the dephosphorylation of its natural substrate 5'-[14C]AMP. We calculated substrate utilization, mean lung transit time (t), and volume of distribution (i.e., central blood volume) of all substrates, as well as inhibitor binding. We also calculated Amax/Km and Bmax products of enzyme mass and kinetic constants for substrates and inhibitor, respectively. As Qb increased, Amax/Km values for all three substrates and Bmax increased linearly, indicating microvascular recruitment. In experiments in which either BPAP and 5'-AMP metabolism or BAGP metabolism and RAC binding were studied concomitantly, a linear relationship was observed between Qb-induced changes in Amax/Km values of BPAP vs 5'-AMP as well as in Amax/Km of BAGP vs Bmax of RAC. Similarly, increasing Qb increased central blood volume and decreased t. Indomethacin had no effect on most of the hemodynamic or enzyme parameters measured. We conclude that in vivo assays of ACE proceed as predicted by Michaelis-Menten kinetics and offer insights into pulmonary endothelial pathophysiology.
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PMID:Assay of pulmonary microvascular endothelial angiotensin-converting enzyme in vivo: comparison of three probes. 829 Oct 66