Andersen K J, McDonald J K
Section for Clinical Research and Molecular Medicine, University of Bergen, Norway.
Am J Physiol. 1987 Oct;253(4):F648-55. doi: 10.1152/ajprenal.1987.253.4.F649.
Differential pelleting of a rat renal cortical homogenate clearly demonstrated the microsomal localization of an N-terminal exopeptidase of the tripeptidyl peptidase (TPP) class that typically requires a free N-terminus to catalyze the release of collagen-related (Gly-Pro-X) "triplets" at pH 7.0 (TPP 7). Once fractionated by differential pelleting, microsomal populations of different size were subfractionated by equilibrium banding in sucrose gradients for the purpose of comparing the distribution profiles and the isopycnic banding densities of TPP 7 to those for known marker enzymes. This analytical approach permitted the localization of these enzymes to specific membrane domains in the renal cortex and provided evidence for the brush-border location of TPP 7. Notably, dipeptidyl peptidase IV (DPP IV), an established plasma membrane exopeptidase with a prolyl-bond specificity, gave banding densities and distributions that were consistent with the presence of both TPP 7 and DPP IV in the same membrane compartment. Because triplets of the Gly-Pro-X type released by TPP 7 would be ideal substrates for DPP IV, a coupled TPP 7-DPP IV exopeptidase mechanism at the luminal surface (brush border) of proximal tubule cells could therefore make a major contribution to the renal degradation and reabsorption of filtered collagen fragments.
对大鼠肾皮质匀浆进行差异离心沉淀，清楚地证明了三肽基肽酶（TPP）类N端外肽酶的微粒体定位，该酶通常需要游离的N端在pH 7.0时催化释放胶原蛋白相关的（甘氨酸-脯氨酸-X）“三联体”（TPP 7）。通过差异离心沉淀进行分级分离后，为了比较TPP 7与已知标记酶的分布图谱和等密度沉降带密度，将不同大小的微粒体群体在蔗糖梯度中通过平衡沉降进一步分级分离。这种分析方法能够将这些酶定位到肾皮质的特定膜结构域，并为TPP 7定位于刷状缘提供了证据。值得注意的是，二肽基肽酶IV（DPP IV）是一种已确定的具有脯氨酰键特异性的质膜外肽酶，其沉降带密度和分布与TPP 7和DPP IV存在于同一膜区室一致。由于TPP 7释放的甘氨酸-脯氨酸-X型三联体将是DPP IV的理想底物，因此近端小管细胞腔表面（刷状缘）的TPP 7-DPP IV外肽酶偶联机制可能对滤过的胶原片段的肾脏降解和重吸收起主要作用。
