Isozyme hybrids within the protruding third loop domain of the barley α‐amylase (β/α)8‐barrel implication for BASI sensitivity and substrate affinity

FEBS Letters

Svensson, Birte²  Chaix, Jean-Claude¹  Guo, Xiao-Jun¹  Rodenburg, Kees W.²  Juge, Nathalie² 

[1] Laboratoire de Biochimie et de Biologie de la Nutrition, CNRS-URA 1820, Faculté des Sciences et Techniques de St-Jérôme, 13 397 Cedex 20 Marseille, France;Carlsberg Laboratory, Department of Chemistry, Gamle Carlsberg Vej 10, DK-2500 Copenhagen Valby, Denmark

关键词: α-Amylase/subtilisin inhibitor;    Domain function;    Isozyme hybrid;    Recombinant α-amylase;    Yeast homeologous;    recombination;    Barley;    AMY1 and AMY2;    barley α-amylase isozymes 1 and 2;    AMY1-(1–112)-AMY2-(112–403);    etc.;    hybrid containing His1-Thr112 of AMY1 and Pro112-Ile403 of AMY2;    etc.;    BASI;    barley α-amylase/subtilisin inhibitor;    pNPG7;    p-nitrophenyl α-d-maltoheptaoside;   

DOI  :  10.1016/0014-5793(95)00291-G

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

Barley α-amylase isozymes AMY1 and AMY2 contain three structural domains: a catalytic (β/α)₈-barrel (domain A) with a protruding loop (domain B; residues 89–152) that binds Ca²⁺, and a small C-terminal domain. Different parts of domain B secure isozyme specific properties as identified for three AMY1–AMY2 hybrids, obtained by homeologous recombination in yeast, with crossing-over at residues 112, 116, and 144. The AMY1 regions Val⁹⁰-Thr¹¹² and Ala¹⁴⁵-Leu¹⁶¹ thus confer high affinities for the substrates p-nitrophenyl α-d-maltoheptaoside and amylose, respectively. Leu¹¹⁷-Phe¹⁴⁴, and to a lesser degree Ala¹⁴⁵-Leu¹⁶¹, are critical for the stability at low pH characteristic of AMY1 and for the sensitivity to barley α-amylase/subtilisin inhibitor specific to AMY2.

【 授权许可】

Unknown