In:
Microbial Physiology, S. Karger AG, Vol. 20, No. 1 ( 2011), p. 29-42
Abstract:
Based on the sequence homology, we have modeled the three-dimensional structure of 〈 i 〉 Bacillus licheniformis 〈 /i 〉 DnaK ( 〈 i 〉 Bl 〈 /i 〉 DnaK), a counterpart of Hsp70, and identified five different amino acids that might be responsible for maintaining ADP-Mg 〈 sup 〉 2+ 〈 /sup 〉 -Pi in the correct configuration at the ATP-binding cleft of the protein. As compared with wild-type 〈 i 〉 Bl 〈 /i 〉 DnaK, site-directed mutant proteins D8A, N13D, E145A, D168A, and T173A had a dramatic reduction in their chaperone activities. Complementation test revealed that the mutant proteins lost completely the ability to rescue the temperature-sensitive growth defect of 〈 i 〉 Escherichia coli 〈 /i 〉 〈 i 〉 dnaK756-ts 〈 /i 〉 . Wild-type 〈 i 〉 Bl 〈 /i 〉 Dnak assisted the refolding of denatured firefly luciferase, whereas a significant decrease in this ability was observed for the mutant proteins. Simultaneous addition of 〈 i 〉 B. licheniformis 〈 /i 〉 DnaJ, 〈 i 〉 Bl 〈 /i 〉 GrpE, and NR-peptide, did not synergistically stimulate the ATPase activity of D8A, E145A, D168A and T173A. Circular dichroism spectra were nearly identical for wild-type and mutant proteins, and they, except D8A, also exhibited a similar sensitivity towards temperature-induced denaturation. These results suggest that the selected residues are critical for the proper function of 〈 i 〉 Bl 〈 /i 〉 DnaK.
Type of Medium:
Online Resource
ISSN:
2673-1665
,
2673-1673
Language:
English
Publisher:
S. Karger AG
Publication Date:
2011
detail.hit.zdb_id:
3042601-7
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