Involvement of the Arg566 residue of Aeromonas sobria serine protease in substrate specificity.
Involvement of the Arg566 residue of Aeromonas sobria serine protease in substrate specificity.
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DOI：
10.1371/journal.pone.0186392
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发表时间：
2017
期刊：
影响因子：
3.7
通讯作者：
Yamanaka H
中科院分区：
文献类型：
--
作者：
Kobayashi H;Otsubo T;Teraoka F;Ikeda K;Seike S;Takahashi E;Okamoto K;Yoshida T;Tsuge H;Yamanaka H
sobria气单胞菌丝氨酸蛋白酶（ASP）是生物体分泌的胞外丝氨酸蛋白酶。在这里，我们通过合成肽和生物蛋白组分鉴定了ASP的氨基酸残基，这些氨基酸残基有助于底物特异性。结果表明，ASP的566位精氨酸残基（Arg-566）位于ASP靠近催化腔入口的额外封闭区域，参与了底物特异性。ASP的Arg-566残基置换点突变为Ala残基（ASP[R566A]），导致对某一底物的蛋白水解效率下降。此外，由于这种点突变，ASP失去了识别主要底物的能力，ASP[R566A]与广泛的合成底物发生反应。Arg-566可能与底物P3位置的氨基酸残基相互作用，这是切割位点上游的第三个氨基酸残基。另一项利用ASP的伴侣蛋白ORF2蛋白的研究进一步表明，Arg-566也可能在与ORF2的相互作用中发挥重要作用。因此我们得出结论，ASP的Arg-566残基可能负责底物的选择。
Aeromonas sobria serine protease (ASP) is an extracellular serine protease secreted by the organism. Here, we identified the amino acid residue of ASP that contributes to substrate specificity by using both synthetic peptides and biological protein components. The results showed that the arginine residue at position 566 (Arg-566) of ASP, which is located in the extra occluding region of ASP close to an entrance of the catalytic cavity, is involved in the substrate specificity. A substitutional point mutation of the Arg-566 residue of ASP to Ala residue (ASP[R566A]) caused a decrease of the proteolytic efficiency for a certain substrate. In addition, ASP lost the ability to recognize the primary substrate by such a point mutation, and ASP[R566A] reacted to a wide range of synthetic substrates. It is likely that Arg-566 causes an interaction with the amino acid residue at position P3 of the substrate, which is the third amino acid residue upstream from the cleavage site. Another study using ORF2 protein, a chaperone protein of ASP, further suggested that Arg-566 could also play an important role in interaction with ORF2. We therefore conclude that the Arg-566 residue of ASP is likely responsible for the selection of substrates.
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影响因子：
2.1
作者：
Kobayashi, H;Takahashi, E;Okamoto, K
通讯作者：
Okamoto, K
影响因子：
4.4
作者：
Nitta, Hidetoshi;Imamura, Takahisa;Baba, Hideo
通讯作者：
Baba, Hideo
影响因子：
2.6
作者：
Okamoto, K;Nomura, T;Fujii, Y
通讯作者：
Fujii, Y
影响因子：
3.7
作者：
Takahashi E;Ozaki H;Fujii Y;Kobayashi H;Yamanaka H;Arimoto S;Negishi T;Okamoto K
通讯作者：
Okamoto K
影响因子：
4.2
作者：
Tsai, Yao-Hung;Huang, Kuo-Chin;Hsu, Robert Wen-Wei
通讯作者：
Hsu, Robert Wen-Wei