Song L, Fricker L D
Department of Molecular Pharmacology, Albert Einstein College of Medicine, Bronx, New York 10461, USA.
J Biol Chem. 1995 Apr 7;270(14):7963-7. doi: 10.1074/jbc.270.14.7963.
Carboxypeptidase E (CPE) is involved with the biosynthesis of numerous peptide hormones and neurotransmitters. Several forms of CPE have been previously detected in neuroendocrine cells, including a form which is soluble at pH 5.5 (S-CPE), a form which can be extracted from membranes with 1 M NaCl at pH 5.5 (M1-CPE), and a form which requires both 1% Triton X-100 and 1 M NaCl for extraction from membranes at pH 5.5 (M2-CPE). Like other peptide processing enzymes, CPE is known to be sorted into peptide-containing secretory vesicles of the regulated pathway. One mechanism that has been proposed to be important for sorting of regulated pathway proteins is Ca2+ and pH-induced aggregation. CPE purified from bovine pituitary membranes aggregates at pH 5.5 when the concentration of CPE is 0.3 micrograms/microliters or higher, but not when the concentration is 0.01 microgram/microliters. Aggregation of CPE is pH-dependent, with very little aggregation occurring at pH 6 or above. At pH 5.0-5.5, the M2 form of CPE shows a greater tendency to aggregate than the other two forms. At pH 6, Ca2+ concentrations from 1-30 mM increase the aggregation of M1- and M2-CPE, but not S-CPE. The aggregation of M2-CPE does not explain the apparent membrane binding of this protein since the aggregate is solubilized by 1% Triton X-100 at pH 5.5 or by pH 6.0, whereas M2-CPE is not extracted from membranes under these conditions. Taken together, these results are consistent with a model in which the decreasing pH and increasing Ca2+ levels in the trans Golgi network induce the aggregation of CPE, which contributes to the sorting of this protein into regulated pathway secretory vesicles.
羧肽酶E（CPE）参与多种肽类激素和神经递质的生物合成。先前已在神经内分泌细胞中检测到几种形式的CPE，包括在pH 5.5时可溶的形式（S-CPE）、在pH 5.5时能用1 M NaCl从膜中提取的形式（M1-CPE），以及在pH 5.5时需要1% Triton X-100和1 M NaCl才能从膜中提取的形式（M2-CPE）。与其他肽加工酶一样，已知CPE会被分选到调节途径中含肽的分泌小泡中。一种被认为对调节途径蛋白分选很重要的机制是Ca2+和pH诱导的聚集。从牛垂体膜中纯化的CPE在浓度为0.3微克/微升或更高时，于pH 5.5会发生聚集，但浓度为0.01微克/微升时则不会。CPE的聚集依赖于pH，在pH 6或更高时几乎不发生聚集。在pH 5.0 - 5.5时，CPE的M2形式比其他两种形式表现出更大的聚集倾向。在pH 6时，1 - 30 mM 的Ca2+浓度会增加M1-和M2-CPE的聚集，但不会增加S-CPE的聚集。M2-CPE的聚集并不能解释该蛋白明显的膜结合情况，因为聚集体在pH 5.5时可被1% Triton X-100溶解，或在pH 6.0时被溶解，而在这些条件下M2-CPE不能从膜中提取出来。综上所述，这些结果与一个模型一致，即反式高尔基体网络中pH降低和Ca2+水平升高会诱导CPE聚集，这有助于将该蛋白分选到调节途径的分泌小泡中。
