A human RNA viral cysteine proteinase that depends upon a unique Zn2+-binding finger connecting the two domains of a papain-like fold .
A human RNA viral cysteine proteinase that depends upon a unique Zn2+-binding finger connecting the two domains of a papain-like fold .
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DOI：
10.1074/jbc.274.21.14918
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发表时间：
1999-05-21
期刊：
影响因子：
--
通讯作者：
Gorbalenya AE
中科院分区：
文献类型：
--
作者：
Herold J;Siddell SG;Gorbalenya AE
人冠状病毒229 E（HCoV）的一种半胱氨酸蛋白酶，木瓜蛋白酶样蛋白酶（PL 1 pro），通过在Gly 111和Asn 112之间的切割来调节复制酶多聚蛋白pp 1a和ppa 1ab的表达，Gly 111和Asn 112位于其自身催化残基Cys 1054的上游。在这份报告中，使用生物信息学工具，我们预测，与其遥远的细胞同源物不同，HCoV PL 1 pro及其冠状病毒亲属具有保守性较差的Zn 2+指，连接木瓜蛋白酶样折叠的左右手结构域。光发射光谱法已被用来确认Zn 2+的存在下，在一个纯化的和蛋白水解活性形式的HCoV PL 1亲融合theEscherichia coli麦芽糖结合蛋白。在使用重组蛋白的变性/复性实验中，其活性被证明是强烈依赖于Zn 2+，这可以部分地被Co 2+在复性过程中取代。重组的含Zn 2+的PL 1 pro对1，10-菲咯啉不敏感，并且Zn 2+耗尽的蛋白质在复性后不会通过添加Zn 2+而重新活化。与所提出的Zn 2+的基本结构作用相一致，PL 1 pro通过Zn 2+指中的突变选择性地失活，包括预测协调Zn 2+的四个保守Cys残基中的任何一个的替换。HCoV PL 1 pro的独特结构域组织为调节过程提供了一个潜在的框架，并可能表明这种酶的非蛋白水解活性。
A cysteine proteinase, papain-like proteinase (PL1pro), of the human coronavirus 229E (HCoV) regulates the expression of the replicase polyproteins, pp1a and ppa1ab, by cleavage between Gly111 and Asn112, far upstream of its own catalytic residue Cys1054. In this report, using bioinformatics tools, we predict that, unlike its distant cellular homologues, HCoV PL1pro and its coronaviral relatives have a poorly conserved Zn2+ finger connecting the left and right hand domains of a papain-like fold. Optical emission spectrometry has been used to confirm the presence of Zn2+ in a purified and proteolytically active form of the HCoV PL1pro fused with theEscherichia coli maltose-binding protein. In denaturation/renaturation experiments using the recombinant protein, its activity was shown to be strongly dependent upon Zn2+, which could be partly substituted by Co2+ during renaturation. The reconstituted, Zn2+-containing PL1pro was not sensitive to 1,10-phenanthroline, and the Zn2+-depleted protein was not reactivated by adding Zn2+ after renaturation. Consistent with the proposed essential structural role of Zn2+, PL1pro was selectively inactivated by mutations in the Zn2+ finger, including replacements of any of four conserved Cys residues predicted to co-ordinate Zn2+. The unique domain organization of HCoV PL1pro provides a potential framework for regulatory processes and may be indicative of a nonproteolytic activity of this enzyme.
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影响因子：
3.5
作者：
Gorbalenya AE;Koonin EV;Lai MM
通讯作者：
Lai MM
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3.5
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GORBALENYA, AE;DONCHENKO, AP;KOONIN, EV
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ALLAIRE, M;CHERNAIA, MM;JAMES, MNG
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5.6
作者：
BAKER, EN
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