Headrick J P, Willis R J
Division of Science and Technology, Griffith University, Nathan, Queensland, Australia.
Biochem J. 1989 Jul 15;261(2):541-50. doi: 10.1042/bj2610541.
Changes in 5'-nucleotidase activity were calculated on the basis of alterations in ATP, ADP, phosphocreatine, Pi, Mg2+, IMP and AMP, determined by using 31P n.m.r. spectroscopy and h.p.l.c., during isoprenaline infusion, graded hypoxia and graded underperfusion in isolated rat heart. Calculated activity changes were compared with the total efflux of purines (adenosine + inosine + hypoxanthine) in order to assess the involvement of various 5'-nucleotidases in formation of adenosine. Purine efflux exhibited an exponential relation with cytosolic [AMP] during isoprenaline infusion and hypoxia (r = 0.92 and 0.95 respectively), supporting allosteric activation of 5'-nucleotidase under these conditions. Purine efflux displayed a linear relation with cytosolic [AMP] during graded ischaemia (r = 0.96), supporting substrate regulation in the ischaemic heart. The calculated activities of membrane-bound ecto-5'-nucleotidase were similar to the observed relations between purine efflux and cytosolic [AMP] in all hearts. The calculated activities of the ATP-activated cytosolic and lysosomal enzymes and of the ATP-inhibited cytosolic 5'-nucleotidase could not explain the observed release of purines under the conditions examined. These results indicate that the kinetic characteristics of the membrane-bound ecto-enzyme are consistent with an important role in the formation of extracellular adenosine, whereas the characteristics of the other 5'-nucleotidases are inconsistent with roles in adenosine formation under the conditions of the present study.
在离体大鼠心脏中，通过31P核磁共振光谱法和高效液相色谱法测定了异丙肾上腺素输注、分级低氧和分级灌注不足期间ATP、ADP、磷酸肌酸、Pi、Mg2+、IMP和AMP的变化，在此基础上计算5'-核苷酸酶活性的变化。将计算得到的活性变化与嘌呤（腺苷+肌苷+次黄嘌呤）的总流出量进行比较，以评估各种5'-核苷酸酶在腺苷形成中的作用。在异丙肾上腺素输注和低氧期间，嘌呤流出量与胞质[AMP]呈指数关系（分别为r = 0.92和0.95），支持在这些条件下5'-核苷酸酶的变构激活。在分级缺血期间，嘌呤流出量与胞质[AMP]呈线性关系（r = 0.96），支持缺血心脏中的底物调节。在所有心脏中，计算得到的膜结合ecto-5'-核苷酸酶活性与观察到的嘌呤流出量和胞质[AMP]之间的关系相似。在研究的条件下，计算得到的ATP激活的胞质和溶酶体酶以及ATP抑制的胞质5'-核苷酸酶的活性无法解释观察到的嘌呤释放。这些结果表明，膜结合ecto-酶的动力学特征与在细胞外腺苷形成中起重要作用一致，而其他5'-核苷酸酶的特征与本研究条件下腺苷形成中的作用不一致。
