The glycosyl hydrolase activity of <i>S. pneumoniae</i> GHIP. NiuSiqiang LuoMiao TangJian ZhouHua ZhangYangli MinXun CaiXuefei ZhangWenlu XuWenchu LiDefeng DingJingjin HuYonglin WangDacheng HuangAilong YinYibin WangDeqiang 2013 <p>(A) <i>S. pneumoniae</i> GHIP (green) superimposed on its homolog from <i>Bacillus anthracis</i>, PlyB (cyan) (PDB code 2NW0). The magenta arrow points toward the major difference, where the N-terminal regions form a helix (α1) which is absent in PlyB. The sticks represent the four key acidic residues to their enzyme activities, inculding Asp56, Asp154, Glu156, and Asp245 of GHIP and Asp6, Asp90, Asp92, and Asp171 of PlyB. (B) The enlarged image of the four key residues at the active site. Close up view of <i>S. pneumoniae</i> GHIP showing overlay of key acidic residues: Asp56, Asp154, Glu156, and Asp245 of GHIP and Asp6, Asp90, Asp92, and Asp171 of PlyB, which exhibit similar locations and orientations. (C) The month of TIM barrel in <i>S. pneumoniae</i> GHIP is active site which contains 14 residues in sticks, including Asp56, Ser58, Ser84, Tyr121, Tyr123, Glu154, Glu156, Asp157, Tyr185, Tyr209, Asp212, Ser233, Asp243 and Asp245. The GHIP structure is shown in the context of a transparent (gray) surface. (D) The enlarged image of the 14 residues at the active site and the background is the electrostatic potential surface of <i>S. pneumoniae</i> GHIP. Saturated red indicates Φ<−10 kiloteslas/e, and saturated blue indicates Φ>10 kiloteslas/e, T = 20°C. (E) & (F) The temperature and pH activity analyses of <i>S. pneumoniae</i> GHIP. Peptidoglycan hydrolase activity was measured at various pH (4.0 to 8.0) and temperatures (25 to 45°C) ranges as described in the Materials and Methods. (G) Hydrolase activity of <i>S. pneumoniae</i> GHIP on PNP-(GlcNAc)<sub>5</sub>. Lane 1 represents the positive control using HEWL (egg white lysozyme); lane 2 is native GHIP; lanes 3–6 are active-site mutants D56A, D154A, E156A, and D245A, respectively. Asterisks denote values significantly different from the wild-type strain by Student’s <i>t</i>-test (*, P<0.05; **, P<0.01).</p>