Partial homology of stress glycoprotein GP62 with HSP70.
Thermotolerance and heat resistance are often associated with elevated levels of heat shock proteins (HSPs) and a selective increase in protein glycosylation. In the present study, we have characterized heat stress-induced protein glycosylation in M21 cells, derived from the rat fibroblast line, Rat-1. M21 cells are characterized by constitutive overexpression of human HSP70 gene and show increased heat resistance without loss of its normal capacity for thermotolerance development after heat conditioning (Li et al., 1991, Proc. Natl. Acad. Sci. USA 88, 1681-1685). The data presented here show that the elevated heat resistance in these cells is associated not only with the constitutive overexpression of human HSP70, but also with increased glycosylation of a major stress glycoprotein, GP62 (Mr of 62,000). We further purified GP62 by sequential preparative isoelectric focusing and two-dimensional isoelectric focusing/SDS-polyacrylamide gel electrophoresis. The purified protein was digested and partially characterized by microsequencing of two peptide fragments, comprising of 14-15 amino acids each. These fragments had a 100% sequence homology with HSP70 and a 71-100% sequence homology with HSC70 from various species. Western blotting using both HSP70 and HSC70 antibodies showed positive reactivity of GP62 with HSP70. Affinity characterizations showed strong binding of GP62 to wheat germ agglutinin and concanavalin A, consistent with the presence of both alpha-D-mannosyl/glucosyl and N-acetyl-beta-D-glucosylaminyl/glucosamine oligomer residues in GP62. These data confirm the glycosylated status of GP62 and indicate that GP62 is a heat stress-induced glycoprotein with partial homology to HSP70.
Jethmalani
S M
SM
Department of Medicine, University of Arkansas for Medical Sciences, Little Rock 72205-5484, USA.
Henle
K J
KJ
D000595:Amino Acid Sequence; D000818:Animals; D005347:Fibroblasts / Q000166:cytology; D006023:Glycoproteins / Q000096:biosynthesis* / Q000145:classification; D018840:HSP70 Heat-Shock Proteins / Q000096:biosynthesis* / Q000737:chemistry; D018869:Heat-Shock Response; D006358:Hot Temperature; D006801:Humans; D008969:Molecular Sequence Data; D051381:Rats; D011994:Recombinant Proteins / Q000096:biosynthesis; D017421:Sequence Analysis; D017386:Sequence Homology, Amino Acid
D006023:Glycoproteins; D018840:HSP70 Heat-Shock Proteins; D011994:Recombinant Proteins