Mutations in the C-terminal fragment of DnaK affecting peptide binding.
Escherichia coli DnaK acts as a molecular chaperone through its ATP-regulated binding and release of polypeptide substrates. Overexpressing a C-terminal fragment (CTF) of DnaK (Gly-384 to Lys-638) containing the polypeptide substrate binding domain is lethal in wild-type E. coli. This dominant-negative phenotype may result from the nonproductive binding of CTF to cellular polypeptide targets of DnaK. Mutations affecting DnaK substrate binding were identified by selecting noncytotoxic CTF mutants followed by in vitro screening. The clustering of such mutations in the three-dimensional structure of CTF suggests the model that loops L1,2 and L4,5 form a rigid core structure critical for interactions with substrate.
Burkholder
W F
WF
Department of Biochemistry and Molecular Biophysics, Columbia University, New York, NY 10032, USA.
Zhao
X
X
Zhu
X
X
Hendrickson
W A
WA
Gragerov
A
A
Gottesman
M E
ME
D001426:Bacterial Proteins / Q000737:chemistry; D029968:Escherichia coli Proteins; D005799:Genes, Dominant; D005816:Genetic Complementation Test; D018840:HSP70 Heat-Shock Proteins / Q000737:chemistry* / Q000235:genetics; D008958:Models, Molecular; D018832:Molecular Chaperones / Q000737:chemistry* / Q000235:genetics; D010446:Peptide Fragments / Q000737:chemistry; D017354:Point Mutation; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins; D013329:Structure-Activity Relationship
D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D018840:HSP70 Heat-Shock Proteins; D018832:Molecular Chaperones; D010446:Peptide Fragments; D011994:Recombinant Proteins; C063842:dnaK protein, E coli