مشخصات پژوهش

tInside the cells, proteins are surrounded by mixtures of different osmolytes. However, our current under-standing of the combinatorial effects of such mixtures on the stability of proteins remains elusive. Inthe present study, the stability and structure of recombinant pyrazinamidase (PZase) from Mycobac-terium tuberculosis were analyzed in the presence of stabilizing osmolytes (sorbitol, sucrose and glycerol)and alcohols (methanol, ethanol, isopropanol and n-propanol). The far-UV and near-UV circular dichro-ism (CD), intrinsic fluorescence and thermostability results indicated that methanol, unexpectedly, hasstronger effect on destabilization of the enzyme compared to ethanol which has larger log P. Interestingly,the relative half-life of PZase was longer in mixtures methanol with the osmolytes, sorbitol or sucrose(expectedly), or glycerol (unexpectedly), compared to other alcohols. Molecular dynamics simulationresults showed that methanol increases the flexibility of region 5-40 and loop 51-71 in the PZase, whichare potentially crucial for the stability and activity of the enzyme, respectively. Our results indicated thatmethanol can interact with PZase via hydrophobic interactions and hydrogen bonds, and therefore result-ing in destabilization of the structure of the enzyme. In addition, glycerol probably increases the stabilityof the enzyme in methanol by disrupting the unfavorable hydrophobic interactions and hydrogen bonds.