Purification of the major mammalian heat shock proteins

• Published in: The Journal of biological chemistry Vol. 257; no. 24; pp. 14949 - 14959
• Main Authors: Welch, W J, Feramisco, J R
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 25-12-1982; American Society for Biochemistry and Molecular Biology
• Subjects: Electrophoresis, Polyacrylamide Gel; Heat-Shock Proteins; HeLa Cells - analysis; Hot Temperature; Humans; Molecular Weight; Proteins - isolation & purification
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1016/S0021-9258(18)33376-3

The major mammalian heat shock or "stress" proteins (molecular masses of 90,000, 72,000, and 73,000 daltons) have been purified from stressed HeLa cells. The 90,000-dalton protein co-purified with small amounts of a 100,000-dalton protein which was identified as one of the other stress proteins in these cells. The 72,000- and 73,000-dalton proteins co-purified throughout the fractionation scheme, apparently as a mixture of monomeric forms of the two proteins. From sedimentation velocity and gel filtration analysis, it was found that the 90,000/100,000-dalton protein mixture had a Stokes radius of 69A and a s20,w value of 5.8 while the 72,000/73,000-dalton protein mixture had a Stokes radius of 42.6A and a s20,w value of 4.3. The purified proteins migrated identically in two-dimensional gel electrophoretograms with their counterparts from total cell lysates of [35S]methionine-labeled stressed HeLa cells. Peptide mapping experiments indicated that the 72,000- and 73,000-dalton proteins contained common peptides while the 90,000- and 100,000-dalton proteins appeared to be distinct. Amino acid analysis of the 90,000- and a mixture of the 72,000/73,000-dalton proteins showed that both contained relatively high amounts of Asp/Asn and Glu/Gln.