title: Solution structure and backbone dynamics of streptopain: insight into diverse substrate specificity.
abstract: Streptococcal pyrogenic exotoxin B (SPE B) is a cysteine protease expressed by Streptococcus pyogenes. The D9N, G163S, G163S/A172S, and G239D mutant proteins were expressed to study the effect of the allelic variants on their protease activity. In contrast to other mutants, the G239D mutant was approximately 12-fold less active. The Gly-239 residue is located within the C-terminal S230-G239 region, which cannot be observed in the x-ray structure. The three-dimensional structure and backbone dynamics of the 28-kDa mature SPE B (mSPE B) were determined. Unlike the x-ray structure of the 40-kDa zymogen SPE B (proSPE B), we observed the interactions between the C-terminal loop and the active site residues in mSPE B. The structural differences between mSPE B and proSPE B were the conformation of the C-terminal loop and the orientation of the catalytic His-195 residue, suggesting that activation and inactivation of SPE B is involved in the His-195 side-chain rotation. Dynamics analysis of mSPE B and the mSPE B/inhibitor complexes showed that the catalytic and C-terminal loops were the most flexible regions with low order parameter values of 0.5 to 0.8 and exhibited the motion on the ps/ns timescale. These findings suggest that the flexible C-terminal loop of SPE B may play an important role in controlling the substrate binding, resulting in its broad substrate specificity.
authors.affiliation: Departments of Biochemistry, Microbiology and Immunology, Medical Technology, and Pediatrics, National Cheng Kung University College of Medicine, 1 University Road, Tainan 701, Taiwan.
authors.forename: Chih-Chieh
authors.initials: CC
authors.lastname: Wang
authors.forename: Hsiang-Chee
authors.initials: HC
authors.lastname: Houng
authors.forename: Chun-Liang
authors.initials: CL
authors.lastname: Chen
authors.forename: Pei-Ju
authors.initials: PJ
authors.lastname: Wang
authors.forename: Chih-Feng
authors.initials: CF
authors.lastname: Kuo
authors.forename: Yee-Shin
authors.initials: YS
authors.lastname: Lin
authors.forename: Jiunn-Jong
authors.initials: JJ
authors.lastname: Wu
authors.forename: Ming T
authors.initials: MT
authors.lastname: Lin
authors.forename: Ching-Chuan
authors.initials: CC
authors.lastname: Liu
authors.forename: Wenya
authors.initials: W
authors.lastname: Huang
authors.forename: Woei-Jer
authors.initials: WJ
authors.lastname: Chuang
mesh_terms: D000595:Amino Acid Sequence; D018360:Crystallography, X-Ray; D003546:Cysteine Endopeptidases / Q000737:chemistry / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D019906:Nuclear Magnetic Resonance, Biomolecular; D017434:Protein Structure, Tertiary; D013297:Streptococcus pyogenes / Q000201:enzymology*; D013379:Substrate Specificity
chemical_list: D003546:Cysteine Endopeptidases; C042477:streptopain