title: Molecular basis for auto- and hetero-catalytic maturation of a thermostable subtilase from thermophilic Bacillus sp. WF146.
abstract: The proform of the WF146 protease, an extracellular subtilase produced by thermophilic Bacillus sp. WF146, matures efficiently at high temperatures. Here we report that the proform, which contains an N-terminal propeptide composed of a core domain (N*) and a linker peptide, is intrinsically able to mature via multiple pathways. One autocatalytic pathway is initiated by cis-processing of N* to generate an autoprocessed complex N*-I(WT), and this step is followed by truncation of the linker peptide and degradation of N*. Another autocatalytic pathway is initiated by trans-processing of the linker peptide followed by degradation of N*. Unlike most reported subtilases, the maturation of the WF146 protease occurs not only autocatalytically but also hetero-catalytically whereby heterogeneous proteases accelerate the maturation of the WF146 protease via trans-processing of the proform and N*-I(WT). Although N* acts as an intramolecular chaperone and an inhibitor of the mature enzyme, the linker peptide is susceptible to proteolysis, allowing the trans-processing reaction to occur auto- and hetero-catalytically. These studies also demonstrate that the WF146 protease undergoes subtle structural adjustments during the maturation process and that the binding of Ca(2+) is required for routing the proform to mature properly at high temperatures. Interestingly, under Ca(2+)-free conditions, the proform is cis-processed into a unique propeptide-intermediate complex (N*-I(E)) capable of re-synthesis of the proform. Based on the basic catalytic principle of serine proteases and these experimental results, a mechanism for the cis-processing/re-synthesis equilibrium of the proform and the role of the linker peptide in regulation of this equilibrium has been proposed.
authors.affiliation: From the State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan 430072, China.
authors.forename: Hui
authors.initials: H
authors.lastname: Zhu
authors.forename: Bi-Lin
authors.initials: BL
authors.lastname: Xu
authors.forename: Xiaoliang
authors.initials: X
authors.lastname: Liang
authors.forename: Yi-Ran
authors.initials: YR
authors.lastname: Yang
authors.forename: Xiao-Feng
authors.initials: XF
authors.lastname: Tang
authors.forename: Bing
authors.initials: B
authors.lastname: Tang
mesh_terms: D000595:Amino Acid Sequence; D001407:Bacillus / Q000201:enzymology*; D002384:Catalysis; D015964:Gene Expression Regulation, Bacterial; D006358:Hot Temperature; D016296:Mutagenesis; D010455:Peptides / Q000737:chemistry; D017510:Protein Folding; D011499:Protein Processing, Post-Translational; D055550:Protein Stability; D057057:Serine Proteases / Q000737:chemistry* / Q000378:metabolism; D013381:Subtilisins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
chemical_list: D010455:Peptides; D057057:Serine Proteases; D013381:Subtilisins
keywords: Bacteria; Enzyme Structure; Maturation; Propeptide; Protein Processing; Protein Stability; Serine Protease; Subtilase