Membrane anchoring and release of carboxypeptidase M: implications for extracellular hydrolysis of peptide hormones.
Carboxypeptidase M (CPM) was discovered as a membrane-bound B-type carboxypeptidase which is widely distributed in a variety of tissues and cells. The amino acid sequence of CPM indicated that the C-terminal hydrophobic region might be a signal for membrane attachment via a glycosylphosphatidylinositol (GPI) anchor. This was demonstrated by [3H)ethanolamine labeling of Madin Darby canine kidney (MDCK) cells which resulted in labeling of the membrane anchor of CPM as shown by immunoprecipitation, polyacrylamide gel electrophoresis and autoradiography. Trypsin released CPM from the membrane, resulting in removal of the radiolabeled ethanolamine. Carboxypeptidase activity was spontaneously and continuously released from MDCK cells into the medium. The released enzyme is a soluble form of CPM as shown by Triton X-114 partitioning, immunoprecipitation, Western blotting, inhibition studies and its neutral pH optimum. CPM was also found in soluble form in biological fluids such as urine and amniotic fluid where it is the primary enzyme that hydrolyzes epidermal growth factor (EGF), producing des-Arg53-EGF. These data indicate that CPM is involved in peptide metabolism on both the cell surface and in extracellular fluids.
Skidgel
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Department of Pharmacology, University of Illinois College of Medicine at Chicago 60612, USA.
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D000595:Amino Acid Sequence; D000653:Amniotic Fluid / Q000737:chemistry; D000818:Animals; D002460:Cell Line; D004285:Dogs; D005110:Extracellular Space / Q000737:chemistry / Q000378:metabolism*; D058851:GPI-Linked Proteins; D006728:Hormones / Q000378:metabolism*; D006868:Hydrolysis; D007668:Kidney / Q000166:cytology; D008565:Membrane Proteins / Q000502:physiology*; D008666:Metalloendopeptidases / Q000032:analysis / Q000378:metabolism*; D008969:Molecular Sequence Data; D010455:Peptides / Q000378:metabolism*; D011485:Protein Binding / Q000502:physiology; D014556:Urine / Q000737:chemistry
D058851:GPI-Linked Proteins; D006728:Hormones; D008565:Membrane Proteins; D010455:Peptides; C058479:carboxypeptidase M; D008666:Metalloendopeptidases