Modulation of Streptomyces Leucine Aminopeptidase by Calcium
Modulation of Streptomyces Leucine Aminopeptidase by Calcium
复制标题
钙对链霉菌亮氨酸氨基肽酶的调节
DOI：
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发表时间：
2006
影响因子：
4.8
通讯作者：
T. Hatanaka
中科院分区：
文献类型：
--
作者：
J. Arima;Yoshiko Uesugi;M. Uraji;S. Yatsushiro;S. Tsuboi;M. Iwabuchi;T. Hatanaka
灰色链霉菌（Streptomyces griseus）亮氨酸氨基肽酶（SGAP）的活性位点含有两个锌原子，作为了解其他金属肽酶的结构和作用机制的模型在临床上具有重要意义。SGAP是一种钙活化和钙稳定酶，其被钙激活与底物特异性相关。在我们之前的研究中，我们发现了一种由链霉菌（Streptomyces sepatus）分泌的非钙调节的亮氨酸氨基肽酶，其初级结构与SGAP具有71%的同源性。在这项研究中，我们利用体内DNA洗牌系统和位点定向诱变的几种突变酶构建了SGAP和S. septatus氨基肽酶的嵌合体，以确定钙调控的关键残基。我们确定了SGAP的关键残基Asp-173和Asp-174与SGAP被钙活化和稳定有关。我们还发现，已知的由SGAP的Asp-3、Ile-4、Asp-262和Asp-266组成的钙结合位点仅有助于钙对SGAP的稳定。此外，我们还发现了一个重要的残基，glu196，它与Asp-173、Asp-174和钙协同作用，以提高SGAP的催化活性。
Streptomyces griseus leucine aminopeptidase (SGAP), which has two zinc atoms in its active site, is clinically important as a model for understanding the structure and mechanism of action of other metallopeptidases. SGAP is a calcium-activated and calcium-stabilized enzyme, and its activation by calcium correlates with substrate specificity. In our previous study, we found a non-calcium-modulated leucine aminopeptidase secreted by Streptomyces septatus, the primary structure of which showed 71% identity with SGAP. In this study, we constructed chimeras of SGAP and S. septatus aminopeptidase by using an in vivo DNA shuffling system and several mutant enzymes by site-directed mutagenesis to identify the key residues in this modulation by calcium. We identified the key residues Asp-173 and Asp-174 of SGAP associated with both SGAP activation and stabilization by calcium. We also showed that the known calcium-binding site, which is composed of Asp-3, Ile-4, Asp-262, and Asp-266 of SGAP, only contributes to SGAP stabilization by calcium. Furthermore, we identified an important residue, Glu-196, that functions in cooperation with Asp-173, Asp-174, and calcium to increase the catalytic activity of SGAP.
影响因子：
2.9
作者：
COMPTON, LA;JOHNSON, WC
通讯作者：
JOHNSON, WC
L-亮氨酸膦酸抑制蛋白水解气单胞菌的氨肽酶。
影响因子：
2.9
作者：
Stamper,C;Bennett,B;Edwards,T;Holz,RC;Ringe,D;Petsko,G
通讯作者：
Petsko,G
DOI：
10.1073/pnas.87.17.6878
发表时间：
1990-09-01
影响因子：
11.1
作者：
BURLEY, SK;DAVID, PR;LIPSCOMB, WN
通讯作者：
LIPSCOMB, WN