ISOZYME HYBRIDS WITHIN THE PROTRUDING 3RD LOOP DOMAIN OF THE BARLEY ALPHA-AMYLASE (BETA/ALPHA)(8)-BARREL IMPLICATION FOR BASI SENSITIVITY AND SUBSTRATE AFFINITY
ISOZYME HYBRIDS WITHIN THE PROTRUDING 3RD LOOP DOMAIN OF THE BARLEY ALPHA-AMYLASE (BETA/ALPHA)(8)-BARREL IMPLICATION FOR BASI SENSITIVITY AND SUBSTRATE AFFINITY
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DOI：
10.1016/0014-5793(95)00291-g
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发表时间：
1995-04-24
期刊：
影响因子：
3.5
通讯作者：
SVENSSON, B
中科院分区：
文献类型：
--
作者：
JUGE, N;RODENBURG, KW;SVENSSON, B
大麦 α-淀粉酶同工酶 AMY1 和 AMY2 包含三个结构域：具有结合 Ca2+ 的突出环（结构域 B；残基 89-152）的催化 (β/α)(8)-桶（结构域 A）和一个小的 C 末端结构域。结构域 B 的不同部分确保了三个 AMY1-AMY2 杂交体的同工酶特异性，这些杂交体是通过酵母中同源重组获得的，在残基 112、116 和 144 处发生交换，AMY1 区域 Val(90)-Thr(112) 和 Ala(145)-Leu(161) 因此赋予底物对硝基苯基的高亲和力分别是α-D-麦芽七苷和直链淀粉。 Leu(117)-Phe(144) 以及较小程度的 Ala(145)-Leu(161) 对于 AMY1 在低 pH 特性下的稳定性以及对 AMY2 特异的大麦 α-淀粉酶/枯草杆菌蛋白酶抑制剂的敏感性至关重要。
Barley alpha-amylase isozymes AMY1 and AMY2 contain three structural domains: a catalytic (beta/alpha)(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 Val(90)-Thr(112) and Ala(145)-Leu(161) thus confer high affinities for the substrates p-nitrophenyl alpha-D-maltoheptaoside and amylose, respectively. Leu(117)-Phe(144), and to a lesser degree Ala(145)-Leu(161), are critical for the stability at low pH characteristic of AMY1 and for the sensitivity to barley alpha-amylase/subtilisin inhibitor specific to AMY2.