Abstract
Mycobacterium leprae HSP18, a major immunodominant antigen of M. leprae pathogen, is a small heat shock protein. Previously, we reported that HSP18 is a molecular chaperone that prevents aggregation of different chemically and thermally stressed client proteins and assists refolding of denatured enzyme at normal temperature. We also demonstrated that it can efficiently prevent the thermal killing of E. coli at higher temperature. However, molecular mechanism behind the chaperone function of HSP18 is still unclear. Therefore, we studied the structure and chaperone function of HSP18 at normal temperature (25°C) as well as at higher temperatures (31–43°C). Our study revealed that the chaperone function of HSP18 is enhanced significantly with increasing temperature. Far- and near-UV CD experiments suggested that its secondary and tertiary structure remain intact in this temperature range (25–43°C). Besides, temperature has no effect on the static oligomeric size of this protein. Subunit exchange study demonstrated that subunits of HSP18 exchange at 25°C with a rate constant of 0.018 min-1. Both rate of subunit exchange and chaperone activity of HSP18 is found to increase with rise in temperature. However, the surface hydrophobicity of HSP18 decreases markedly upon heating and has no correlation with its chaperone function in this temperature range. Furthermore, we observed that HSP18 exhibits diminished chaperone function in the presence of NaCl at 25°C. At elevated temperatures, weakening of interactions between HSP18 and stressed client proteins in the presence of NaCl results in greater reduction of its chaperone function. The oligomeric size, rate of subunit exchange and structural stability of HSP18 were also found to decrease when electrostatic interactions were weakened. These results clearly indicated that subunit exchange and electrostatic interactions play a major role in the chaperone function of HSP18.
Highlights
Exposure to temperatures beyond normal conditions or other stress factors may be lethal to cells
Only 18 kDa antigen has been identified as small heat shock protein and termed as HSP18
Cole et al reported the comparative analysis of M. leprae genome sequence with that of M. tuberculosis and revealed that, despite a massive gene decay in M. leprae, HSP18 gene has been retained [44], hinting over the fact that this small heat shock protein may be critical for virulence and survival of this pathogen
Summary
Exposure to temperatures beyond normal conditions or other stress factors may be lethal to cells. Charge-charge/electrostatic interactions are key to the substrate binding properties of these sHSPs. to understand whether electrostatic interactions play a role in the chaperone function of M. leprae HSP18, we investigated the chaperone activity of HSP18 in the absence or presence of NaCl (0.05–0.5M) using DTT induced aggregation of insulin at 25°C.
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