Variants of the industrially relevant protease KP-43 with suppressed activity under alkaline conditions developed using expanded genetic codes
Variants of the industrially relevant protease KP-43 with suppressed activity under alkaline conditions developed using expanded genetic codes
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使用扩展遗传密码开发的工业相关蛋白酶 KP-43 的变体，在碱性条件下活性受到抑制
DOI：
10.1016/j.bbrep.2018.12.001
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发表时间：
2019
影响因子：
2.7
通讯作者：
Takimura Yasushi
中科院分区：
文献类型：
--
作者：
Osamura Tatsuya;Okuda Mitsuyoshi;Yamaguchi Atsushi;Ohtake Kazumasa;Sakamoto Kensaku;Takimura Yasushi
在本研究中，我们试图通过在催化位点和附近加入3-硝基酪氨酸和3-氯基酪氨酸来控制工业上相关的碱性蛋白酶KP-43的催化活性的pH值。在大肠杆菌宿主细胞中合成了30个在特定位置含有这些非天然氨基酸的KP-43变体，并扩增了遗传密码。在靠近底物结合位点的205位含有3-硝基酪氨酸的变体在中性pH下保持其催化活性，在pH 10.5下活性降低60%。这种碱性结构域的减少对于增强液体洗衣液中酶的稳定性是理想的，而野生型分子对相同的pH值变化的响应增加了20%。在碱性条件下，3-硝基酪氨酸与底物的亲和力降低，这可能是由于3-硝基酪氨酸的酚类pkk低于酪氨酸的酚类pkk而带负电荷。
In the present study, we attempted to control the pH profile of the catalytic activity of the industrially relevant alkaline protease KP-43, by incorporating 3-nitro-l-tyrosine and 3-chloro-l-tyrosine at and near the catalytic site. Thirty KP-43 variants containing these non-natural amino acids at the specific positions were synthesized inEscherichia colihost cells with expanded genetic codes. The variant with 3-nitrotyrosine at position 205, near the substrate binding site, retained its catalytic activity at the neutral pH and showed a 60% activity reduction at pH 10.5. This reduction in the alkaline domain is desirable for enhancing the stability of the enzyme in the liquid laundary detergent, whereas the wild-type molecule showed a 20% increase in response to the same pH shift. The engineered pH dependency of the activity of the variant was ascribed partly to a lowered substrate affinity under the alkaline conditions, in which the incorporated 3-nitrotyrosine was probably charged negatively due to the phenolic pKalower than that of tyrosine.
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影响因子：
14.9
作者：
Mukai T;Hayashi A;Iraha F;Sato A;Ohtake K;Yokoyama S;Sakamoto K
通讯作者：
Sakamoto K
洗涤剂酶：挑战！
DOI：
10.1201/noe1420091625-25
发表时间：
1999
期刊：
Surfactant science series
影响因子：
--
作者：
A. Crutzen;M. L. Douglass
通讯作者：
M. L. Douglass
DOI：
--
发表时间：
2014
期刊：
Research journal of pharmaceutical, biological and chemical sciences
影响因子：
--
作者：
Bashir Sajo Mienda;A. Yahya;Ibrahim A. Galadima;M. S. Shamsir
通讯作者：
M. S. Shamsir
影响因子：
4.8
作者：
Nonaka, T;Fujihashi, M;Miki, K
通讯作者：
Miki, K