Sequential formation of two branched intermediates during protein splicing of class three inteins

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• 作者：Kazuo Tori ; Francine Perler
• 关键词：Intein ; Protein splicing ; Branched intermediate ; Enzyme mechanism ; Cysteine
• 刊名：Extremophiles
• 出版年：2017
• 出版时间：January 2017
• 年：2017
• 卷：21
• 期：1
• 页码：41-49
• 全文大小：958KB
• 刊物类别：Biomedical and Life Sciences
• 刊物主题：Microbiology; Biotechnology; Biochemistry, general; Microbial Ecology;
• 出版者：Springer Japan
• ISSN：1433-4909
• 卷排序：21

文摘

Inteins are the protein equivalent of introns. They are seamlessly removed during post-translational maturation of their host protein (extein). Inteins from extremophiles played a key role in understanding intein-mediated protein splicing. There are currently three classes of inteins defined by catalytic mechanism and sequence signatures. This study demonstrates splicing of three class 3 mini-inteins: Burkholderia vietnamiensis G4 Bvi IcmO intein, Mycobacterium smegmatis MC2 155 Msm DnaB-1 intein and Mycobacterium leprae strain TN Mle DnaB intein. B. vietnamiensis has a broad ecological range and remediates trichloroethene. M. smegmatis is a biofilm forming soil bacteria. Although other intein classes have only a single branched intermediate at the C-terminal splice junction, the class 3 intein reaction pathway includes two branched intermediates. The class 3 specific branched intermediate is formed by an internal cysteine, while the C-terminal branch intermediate is at a serine or threonine in all class 3 inteins except the Bvi IcmO intein, where it is a cysteine. This latter cysteine was unable to compensate for mutation of the class 3-specific internal catalytic cysteine despite the Bvi IcmO intein having an N-terminal splice junction naturally tuned for a cysteine nucleophile, demonstrating the mandatory order of branch intermediates in class 3 inteins.