decoding the folding of burkholderia glumae lipase folding intermediates en route to kinetic stability解码的折叠glumae脂肪酶折叠中间体途中动力学稳定性.pdfVIP

Decoding the Folding of Burkholderia glumae Lipase: Folding Intermediates En Route to Kinetic Stability 1,2 3¤ 4 1,5 Kris Pauwels *, Manuel M. Sanchez del Pino , Georges Feller , Patrick Van Gelder 1 Department of Structural Biology, VIB and Vrije Universiteit Brussel, Brussels, Belgium, 2 National Institute for Medical Research, Molecular Structure Division, London, ´ ´ ´ United Kingdom, 3 Centro de Investigacion Prıncipe Felipe, Laboratorio de Proteomica, Valencia, Spain, 4 Laboratory of Biochemistry, Center for Protein Engineering, ` ` University of Liege, Liege-Sart Tilman, Belgium, 5 L-ProBE, Unit for Structural Biology, Ghent University, Ghent, Belgium Abstract The lipase produced by Burkholderia glumae folds spontaneously into an inactive near-native state and requires a periplasmic chaperone to reach its final active and secretion-competent fold. The B. glumae lipase-specific foldase (Lif) is classified as a member of the steric-chaperone family of which the propeptides of a-lytic protease and subtilisin are the best known representatives. Steric chaperones play a key role in conferring kinetic stability to proteins. However, until present there was no solid experimental evidence that Lif-dependent lipases are kinetically trapped enzymes. By combining thermal denaturation studies with proteolytic resistance experiments and the description of distinct folding intermediates, we demonstrate that the native lipase has a kinetically stable conformation. We