Purification and properties of the Escherichia coli heat shock protein, HtpG.
As a preliminary to the understanding of the function of the highly conserved Escherichia coli heat shock protein HtpG, the protein was purified and partially characterized. The htpG gene was subcloned into the inducible expression vector, pT7-6. Upon induction, the HtpG protein accumulated to approximately 30% of the total protein in the cell. A purification scheme was devised which involved column chromatography on DEAE-cellulose, hydroxylapatite, and Sephacryl S-200. The amino acid composition of the purified protein corresponded closely with the predicted amino acid composition derived from the DNA sequence, and the sequence of the 8 amino-terminal residues matched the predicted sequence exactly. The molecular weight of the denatured protein is 65,500 and the native molecular weight is 144,620, as calculated by using both the Stokes radius and the sedimentation coefficient. As the molecular weight predicted from the DNA sequence is 71,429, this indicates the HtpG protein is a dimer. The HtpG protein was found to be a phosphoprotein. Thus, HtpG is structurally similar to its eukaryotic homologue, hsp83, which is also a phosphoprotein and a dimer.
Spence
J
J
Department of Cellular, Viral, and Molecular Biology, University of Utah Medical Center, Salt Lake City 84132.
Georgopoulos
C
C
D000596:Amino Acids / Q000032:analysis; D001426:Bacterial Proteins; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D002845:Chromatography; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000032:analysis* / Q000235:genetics; D029968:Escherichia coli Proteins; D005822:Genetic Vectors; D018841:HSP90 Heat-Shock Proteins; D006360:Heat-Shock Proteins / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification*; D007163:Immunosorbent Techniques; D008970:Molecular Weight; D010750:Phosphoproteins; D010957:Plasmids; D011401:Promoter Regions, Genetic
D000596:Amino Acids; D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D018841:HSP90 Heat-Shock Proteins; D006360:Heat-Shock Proteins; C489352:HtpG protein, E coli; D010750:Phosphoproteins; C066720:HtpG protein, bacteria