Structural characterization of adrenal chromogranin A and parathyroid secretory protein-I as homologs.
We have isolated and purified adrenal chromogranin A (Ch A) for the purpose of making structural comparisons to parathyroid secretory protein-I (SP-I), because our earlier data indicated these two molecules may be the same protein. An improved purification step, using high-performance liquid chromatography (HPLC), has enabled us to demonstrate that both SP-I and Ch A consists of two species, one of approximately 72,000 Da and one of approximately 66,000 Da. The amino acid composition is the same for all four species. The difference in molecular mass is assumed to be due to carbohydrate content. Cyanogen bromide digestion of each of the four samples, followed by HPLC separation of the generated peptides, resulted in a chromatographic profile that was the same for each digest. Amino acid analysis of the eight peptide fragments obtained from each digest indicates that both species of Ch A and both species of SP-I yielded the same peptide mixtures following this cleavage reaction. One large (approximately 50,000 Da) CNBr peptide was obtained and seven smaller ones, one of which contains cysteine. The large fragment behaved similarly to the intact molecule in a radioimmunoassay. HPLC separation of tryptic digests of Ch A (72,000 Da) and SP-I (72,000 Da) also resulted in elution profiles that were very similar to each other. Amino acid analysis revealed 23 peptides common to each digest. Ch A contained four peptides ranging in size from 4 to 30 residues that were not observed in the SP-I digest. SP-I contained two peptides, each with about 30 residues, that were not found in the Ch A digest. Nothing unusual was noted in any of the uncommon peptides. Thus, both a chemical and an enzymatic digestion of these molecules followed by analysis of the peptides generated, indicates that SP-I and Ch A are nearly identical homologs.
D000311:Adrenal Glands / Q000032:analysis; D000596:Amino Acids / Q000032:analysis; D000818:Animals; D002135:Calcium-Binding Proteins / Q000302:isolation & purification; D002417:Cattle; D055598:Chemical Phenomena; D002621:Chemistry; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange; D053379:Chromogranin A; D002864:Chromogranins / Q000302:isolation & purification; D009419:Nerve Tissue Proteins / Q000302:isolation & purification
D000596:Amino Acids; D002135:Calcium-Binding Proteins; D053379:Chromogranin A; D002864:Chromogranins; D009419:Nerve Tissue Proteins