Chaperone-Like Activity of Mycobacterium tuberculosis Hsp16.3 Does Not Require Its Intact (Native) Structures

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Xiaoyou Chen¹, Xinmiao Fu^2,3, Yu Ma¹, and Zengyi Chang^2,3*

¹Beijing Tuberculosis and Thoracic Tumor Institute, Beijing 101149, China

²State Key Laboratory of Protein Engineering and Plant Genetic Engineering, Peking University, Beijing 100871, China

³College of Life Science, Peking University, Beijing 100871, China; fax: 86-10-6275-1526; E-mail: changzy@pku.edu.cn

^* To whom correspondence should be addressed.

Received August 14, 2004; Revision received September 25, 2004

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Small heat shock proteins (sHsps) were found to exhibit efficient chaperone-like activities under stress conditions although their native structures are severely disturbed. Here, using an alternative approach (site-directed mutagenesis), we obtained two structurally and functionally distinct Mycobacterium tuberculosis Hsp16.3 single-site mutant proteins. The G59W mutant protein (with Gly59 substituted by Trp) is capable of exhibiting efficient chaperone-like activity even under non-stress conditions although its secondary, tertiary, and quaternary structures are very different from that of the wild type protein. By contrast, the G59A mutant protein (with Gly59 substituted by Ala) resembles with the wild type protein in structure and function. These observations suggest that the Gly59 of the Hsp16.3 protein is critical for its folding and assembly. In particular, we propose that the exhibition of chaperone-like activity for Hsp16.3 does not require its intact (native) structures but requires the disturbance of its native structures (i.e., the native structure-disturbed Hsp16.3 retains its chaperone-like activity or even becomes more active). In addition, the behavior of such an active mutant protein (G59W) also strongly supports our previous suggestion that Hsp16.3 exhibits chaperone-like activity via oligomeric dissociation.

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KEY WORDS: chaperone, small heat shock protein, Hsp16.3, oligomer, Mycobacterium tuberculosis

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