HEAT SHOCK-INDUCED PROMPT GLYCOSYLATION - IDENTIFICATION OF P-SG67 ASCALRETICULIN

Acute heat shock initiates the phenomenon of ''prompt'' glycosylation, which is characterized by selective glycosylation of specific cellula r proteins called the prompt stress glycoproteins (P-SG). Prompt glyco sylation rapidly occurs even during short heating periods, e.g. 10 min at 45 degrees C, and is not affected by the presence of cycloheximide (Henle, K. J., Kaushal, G. P., Nagle, W. A., and Nolen, G. T. (1993) Exp. Cell Res. 207, 245-251). The major P-SG in Chinese hamster ovary cells, P-SG67, was characterized by an M(r) of 67,000 and a pI = 5.1. In the present study, we purified P-SG67 by sequential gel fil tration , anion exchange, affinity chromatography with concanavalin A-Sepharos e, and two dimensional isoelectric focusing/SDS-polyacrylamide gel ele ctrophoresis. The purified protein was digested and partially characte rized by microsequencing of three major peptide fragments. The fragmen ts, comprising a total of 46 amino acid residues, had an almost 100% s equence homology with calreticulin and partial homology with calnexin. Calcium binding studies with Ca-45(2+) overlay confirmed that P-SG67 is a Ca2+-binding protein. These observations support the notion that P-SG67 is identical to calreticulin and that the glycosylation status of calreticulin can respond to environmental stress conditions.