Identification of key amino acid residues in Neisseria polysaccharea amylosucrase

FEBS Letters

Svensson, Birte²  Willemot, René-Marc¹  Remaud-Simeon, Magali¹  Monsan, Pierre¹  Potocki de Montalk, Gabrielle¹  Sarçabal, Patricia¹ 

[1] Centre de Bioingénierie Gilbert Durand, UMR CNRS 5504, UR INRA 792, INSA, 135 Avenue de Rangueil, 31 077 Toulouse Cedex 4, France;Department of Chemistry, Carlsberg Laboratory, Gamle Carlsberg Vej 10, DK-2500 Copenhagen Valby, Denmark

关键词: Amylosucrase;    Site-directed mutagenesis;    Active site;    Catalytic nucleophile;    General acid catalyst;    AS;    amylosucrase;    GST;    glutathione S-transferase;    TAA;    Taka-amylase from Aspergillus oryzae;    SDS–PAGE;    sodium dodecylsulfate–polyacrylamide gel electrophoresis;   

DOI  :  10.1016/S0014-5793(00)01567-2

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

Amylosucrase from Neisseria polysaccharea catalyzes the synthesis of an amylose-like polymer from sucrose. Sequence alignment revealed that it belongs to the glycoside hydrolase family 13. Site-directed mutagenesis enabled the identification of functionally important amino acid residues located at the active center. Asp-294 is proposed to act as the catalytic nucleophile and Glu-336 as general acid base catalyst in amylosucrase. The conserved Asp-401, His-195 and His-400 residues are critical for the enzymatic activity. These results provide strong support for the predicted close structural and functional relationship between the sucrose-glucosyltransferases and enzymes of the α-amylase family.

【 授权许可】

Unknown