Chromogranin A: posttranslational modifications in secretory granules.
The primary structure of chromogranin A indicates multiple domains which might be subject to posttranslational modification. We explored chromogranin A's proteolytic cleavage, glycosylation, and possible intermolecular disulfide links, using biochemical and cell biological approaches. Anti-chromogranin A region-specific immunoblots on chromaffin granules suggested bidirectional endoproteolytic cleavage of chromogranin A; control experiments ruled out artifactual cleavage during granule isolation or lysis. Isolation of chromogranin A-derived peptides by gel filtration chromatography or sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), followed by N-terminal amino acid sequencing, established several cleavage sites, including at least two at dibasic sites. Secretion of chromogranin A from bovine chromaffin cells did not initiate further cleavage, nor did prolonged exposure of secreted chromogranins to the secretory cells. The chromogranin A cleavage pattern was qualitatively similar in other neuroendocrine tissues, though cleavage was more complete in adrenal medullary than in anterior pituitary hormone storage vesicles, and N-terminal fragments of 45 and 55 kilodaltons were more prominent in the hypothalamus. A similar cleavage pattern was seen in human pheochromocytoma granules, as judged by chromogranin A region-specific immunoblots, fragment isolation by SDS-PAGE, and microsequencing. The presence of full-length chromogranin A as the core protein of a chromaffin granule soluble proteoglycan was suggested in bovine (but not human) chromaffin granules by glycoprotein staining, chondroitinase ABC digestion, chemical deglycosylation, and region-specific immunoblotting. Human (but not bovine) chromogranin A displayed intermolecular disulfide crosslinks on SDS-PAGE gels and immunoblotting. These results document diverse structural paths that the chromogranin A molecule may take in endocrine secretory cells after its translation.
Barbosa
J A
JA
Department of Medicine, University of California, San Diego.
Gill
B M
BM
Takiyyuddin
M A
MA
O'Connor
D T
DT
D000310:Adrenal Gland Neoplasms / Q000378:metabolism*; D000313:Adrenal Medulla / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D002478:Cells, Cultured; D002837:Chromaffin Granules / Q000378:metabolism*; D053379:Chromogranin A; D002864:Chromogranins / Q000235:genetics* / Q000302:isolation & purification; D015180:Electrophoresis, Gel, Two-Dimensional; D006031:Glycosylation; D006801:Humans; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010446:Peptide Fragments / Q000302:isolation & purification; D010673:Pheochromocytoma / Q000378:metabolism*; D011499:Protein Processing, Post-Translational
C506150:CHGA protein, human; D053379:Chromogranin A; D002864:Chromogranins; D010446:Peptide Fragments