PURIFICATION AND IDENTIFICATION OF A 28-KDA CALCIUM-REGULATED HEAT-STABLE PROTEIN - A NOVEL SECRETAGOGUE-REGULATED PHOSPHOPROTEIN IN EXOCRINE PANCREAS

This study reports the purification and identification of a novel 28 k Da phosphoprotein from rat pancreatic acini, previously described as b eing highly regulated by calcium mobilizing secretagogues, which we ha ve designated calcium-regulated heat-stable protein 28 (CRHSP-28). Int ernal amino acid sequences of purified CRHSP-28 were obtained followin g trypsin digestion and found to match with >95% identity the predicte d amino acid sequence of a novel cDNA recently identified as being hig hly expressed in human breast carcinomas, Verification that this cDNA codes for human CRHSP-28 was demonstrated by the ability of antiserum raised against purified rat CRHSP-28 to recognize the recombinant huma n protein when expressed in bacteria. Furthermore, this antibody was f ound to specifically react with CRHSP-28 in rat acini following one- a nd tno dimensional electrophoresis and underwent a marked acidic shift in mobility after cholecystokinin stimulation, a phenomenon indicativ e of an increase in its phosphorylation. CRHSP-28 is predicted to be e xtremely hydrophilic, is phosphorylated entirely on serine residues, a nd bears little homology to any known proteins, Finally, the distribut ion of the CRHSP-28 protein in various rat tissues revealed that altho ugh it was present at low levels in almost all tissues, it was most hi ghly expressed in pancreas, followed by the gastric, intestinal, and c olonic mucosa. In view of its relative abundance throughout the digest ive system and its apparent regulation by calcium-mobilizing agents, t his protein may provide valuable insight into the mechanism(s) of calc ium signaling in these tissues.