Calcium- and pH-dependent aggregation of carboxypeptidase E.
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.
Song
L
L
Department of Molecular Pharmacology, Albert Einstein College of Medicine, Bronx, New York 10461, USA.
Fricker
L D
LD
D000595:Amino Acid Sequence; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D043423:Carboxypeptidase H; D002268:Carboxypeptidases / Q000302:isolation & purification / Q000378:metabolism*; D002417:Cattle; D004591:Electrophoresis, Polyacrylamide Gel; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D010902:Pituitary Gland / Q000201:enzymology
D002268:Carboxypeptidases; D043423:Carboxypeptidase H; D002118:Calcium