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首页> 外文期刊>Journal of Molecular Biology >FUNCTIONAL COOPERATION OF THE MITOCHONDRIAL PROCESSING PEPTIDASE SUBUNITS
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FUNCTIONAL COOPERATION OF THE MITOCHONDRIAL PROCESSING PEPTIDASE SUBUNITS

机译：线粒体加工肽酶亚基的功能合作

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Domains important for the activity of the heterodimeric mitochondrial processing peptidase (MPP) were investigated, by inserting one alanine residue at ten positions along the polypeptide chain of the beta-subunit (beta-MPP). An alanine residue inserted after Glu70, Ser114, Lys215 and Ser314 respectively, abolished the cleavage activity of MPP. When the alpha-subunit (alpha-MPP) was co-expressed with N-terminal hexa-histidine tagged beta-MPP, alpha-MPP was co-eluted from a nickel-derivatized affinity resin, with a 1:1 stochiometry, both with wild-type beta-MPP and with the mutants with alanine inserted after Ser114 and Ser314. The mutants with alanine inserted after Glu70 and Lys215 did not associate with alpha-MPP. The mutagenesis studies indicate that: (1) the whole HXXEHX76H region of beta-MPP is important for the proper conformation of the active site of MPP and may also be in contact with alpha-MPP; (2) the non-conserved central region surrounding Lys215 is involved in the interaction with alpha-MPP; and (3) the carboxy-terminal region of beta-MPP surrounding Ser314 is also of importance for the catalysis. Cross-linking studies indicated that purified alpha-MPP bound a precursor protein in the absence of any beta-MPP. Futhermore, the interaction of MPP and its subunits with a peptide substrate, as analyzed by surface plasmon resonance, showed that alpha-MPP bound a peptide substrate as efficiently as MPP. The data suggest that the alpha-subunit is responsible for the binding of mitochondrial presequences prior their presentation to the catalytic site of MPP. (C) 1997 Academic Press Limited. [References: 34]
机译：通过在沿β-亚基（β-MPP）多肽链的十个位置插入一个丙氨酸残基，研究了对异源二聚体线粒体加工肽酶（MPP）的活性重要的域。分别插入Glu70，Ser114，Lys215和Ser314之后的丙氨酸残基消除了MPP的切割活性。当α亚基（alpha-MPP）与N末端六组氨酸标记的beta-MPP共表达时，alpha-MPP从镍衍生的亲和树脂中以1：1的化学计量共洗脱，野生型β-MPP，以及在Ser114和Ser314之后插入了带有丙氨酸的突变体。在Glu70和Lys215之后插入了丙氨酸的突变体与alpha-MPP不相关。诱变研究表明：（1）β-MPP的整个HXXEHX76H区对于正确配置MPP的活性位点很重要，也可能与α-MPP接触; （2）Lys215周围的非保守中心区域与α-MPP相互作用。 （3）围绕Ser314的β-MPP的羧基末端区域对于催化也很重要。交联研究表明，在不存在任何β-MPP的情况下，纯化的α-MPP会结合前体蛋白。此外，通过表面等离子体共振分析，MPP及其亚基与肽底物的相互作用表明，α-MPP与MPP一样有效地结合肽底物。数据表明，α-亚基负责线粒体先序列的结合，然后将其呈递至MPP的催化位点。 （C）1997 Academic Press Limited。 [参考：34]

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