Artificial chaperone mediated refolding of xylanase from an alkalophilic thermophilic Bacillus sp.

摘要: To gain insight into the molecular aspects of unfolding/refolding enzymes from extremophilic organisms, we have used xylanase an alkalophilic thermophilic Bacillus as model system. Kinetics denaturation/renaturation were monitored using intrinsic fluorescence studies. The protein measurements suggested a putative intermediate state present in 0.08 M guanidine hydrochloride with emission maximum 345 nm; far-UV circular dichroism spectra revealed content secondary structure similar to native enzyme. Studies fluorescent apolar probe 1-anilinonapthalene-8-sulfonate (1,8-ANS) consistent presence increased hydrophobic surfaces compared or fully unfolded protein. refolding Xyl II, was attempted by relatively new strategy artificial chaperone assisted two-step method. found bind detergent transiently and subsequent addition methyl-beta-cyclodextrin helped strip assist folding. Our findings that stabilized folding pathway seemingly equivalent described molten globule. reactivation II affected ionic well nonionic detergents. However, cationic cetyltrimethylammonium bromide (CTAB) provided (threefold) 'delayed addition' experiments acts suppressing initial aggregate formation not dissolving aggregates. relevance our role chaperones vivo is discussed.