Cloning and characterization of a haloarchaeal heat shock protein 70 functionally expressed in Escherichia coli.
The Hsp70 molecular chaperone machine is constituted by the 70-kDa heat shock protein Hsp70 (DnaK), cochaperone protein Hsp40 (DnaJ) and a nucleotide-exchange factor GrpE. Although it is one of the best-characterized molecular chaperone machines, little is known about it in archaea. A 5.2-kb region containing the hsp70 (dnaK) gene was cloned from Natrinema sp. J7 strain and sequenced. It contained the Hsp70 chaperone machine gene locus arranged unidirectionally in the order of grpE, hsp70 and hsp40 (dnaJ). The hsp70 gene from Natrinema sp. J7 was overexpressed in Escherichia coli BL21 (DE3). The recombinant Hsp70 protein was in a soluble and active form, and its ATPase activity was optimally active in 2.0 M KCl, whereas NaCl had less effect. In vivo, the haloarchaeal hsp70 gene allowed an E. coli dnak-null mutant to propagate lambda phages and grow at 42 degrees C. The results suggested that haloarchaeal Hsp70 should be beneficial for extreme halophiles survival in low-salt environments.
Zhang
Hao
H
Laboratory of Microbial Genetics, College of Life Sciences, Wuhan University, Wuhan, China.
Lin
Lu
L
Zeng
Chi
C
Shen
Ping
P
Huang
Yu-Ping
YP
D000222:Adaptation, Physiological / Q000235:genetics; D000251:Adenosine Triphosphatases / Q000737:chemistry / Q000235:genetics / Q000502:physiology; D019843:Archaeal Proteins / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D001483:Base Sequence; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D029968:Escherichia coli Proteins / Q000235:genetics; D019766:Genes, Archaeal; D005816:Genetic Complementation Test; D018840:HSP70 Heat-Shock Proteins / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D006216:Halobacteriaceae / Q000235:genetics / Q000502:physiology*; D006358:Hot Temperature; D008969:Molecular Sequence Data; D009995:Osmosis; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D024363:Transcription Initiation Site
D019843:Archaeal Proteins; D029968:Escherichia coli Proteins; D018840:HSP70 Heat-Shock Proteins; D011994:Recombinant Proteins; D000251:Adenosine Triphosphatases; C063842:dnaK protein, E coli