Laboratório de Estudos sobre o Sistema Purinérgico, Departamento de Bioquímica, Instituto de Ciências Básicas da Saúde, Universidade Federal do Rio Grande do Sul, Porto Alegre, RS, Brazil.
J Enzyme Inhib Med Chem. 2012 Feb;27(1):29-36. doi: 10.3109/14756366.2011.574129. Epub 2011 May 3.
In this study, we have reported the kinetic and biochemical characterization of ectonucleotide pyrophosphatase/phosphodiesterase (E-NPP) activity in rat cardiac fractions, one soluble and the other enriched in vesicles derived from sarcoplasmic reticulum. Both fractions demonstrated E-NPP activities, which could be observed by extracellular hydrolysis of p-nitrophenyl-5'-thymidine monophosphate (p-Nph-5'-TMP) and other biochemical characteristics. The K(M) values for the hydrolysis of p-Nph-5'-TMP in soluble and microsomal fractions were 118.53 ± 27.28 and 91.92 ± 12.49 µM, respectively. The V(max) values calculated were 2.56 ± 0.15 and 113.87 ± 21.09 nmol p-nitrophenol/min/mg of protein in soluble and microsomal fractions, respectively. Among the compounds tested to evaluate the possible activity of other enzymes on p-Nph-5'-TMP hydrolysis, only suramin (0.25 mM) produced a significant inhibition of substrate hydrolysis. Thus, our results strongly suggest the presence of E-NPP enzymes in subcellular fractions of rat heart, which could be involved in nucleotide signalling in the cardiac tissue.
在这项研究中，我们报告了大鼠心脏部分（一种可溶性部分和另一种富含肌浆网衍生小泡的部分）中核苷酸焦磷酸酶/磷酸二酯酶（E-NPP）活性的动力学和生化特征。这两个部分都表现出 E-NPP 活性，可以通过细胞外水解 p-硝基苯-5'-胸苷单磷酸（p-Nph-5'-TMP）和其他生化特征来观察到。可溶性和微粒体部分水解 p-Nph-5'-TMP 的 K(m)值分别为 118.53 ± 27.28 和 91.92 ± 12.49 µM。计算得出的 V(max)值分别为 2.56 ± 0.15 和 113.87 ± 21.09 nmol p-硝基苯酚/分钟/毫克蛋白。在所测试的化合物中，只有苏拉明（0.25 mM）对底物水解产生了显著抑制，从而强烈提示大鼠心脏亚细胞部分存在 E-NPP 酶，这些酶可能参与心脏组织中的核苷酸信号转导。
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