Spatial orientation of mitochondrial processing peptidase and a preprotein revealed by fluorescence resonance energy transfer

Tomonori G. Nishino, Ken Kitano, Katsuhiko Kojima, Tadashi Ogishima, Akio Ito, Sakae Kitada

研究成果: Contribution to journalArticle査読

4 被引用数 (Scopus)

抄録

Mitochondrial processing peptidase (MPP), which is composed of heterodimeric α-MPP and β-MPP subunits. It specifically recognizes mitochondrial preproteins and removes their basic N-terminal signal prepeptides. In order to elucidate the spatial orientation of the preproteins toward MPP, which has been missed by crystal structures of a yeast MPP including a synthetic prepeptide in its acidic proteolytic chamber, we analysed the fluorescence resonance energy transfer (FRET) between EGFP fused to a yeast aconitase presequence (preEGFP) and regiospecific 7-dietylamino-3-(4′-maleimidyl phenyl)-4-methyl coumarin (CPM)-labelled yeast MPPs. FRET efficiencies of 65 and 55% were observed between the EGFP chromophore and CPM-Ser84 and -Lys156 of β-MPP, respectively, leading to calculated distances between the molecules of 48 and 50 Å, respectively. Considering the FRET results and the structural validity based on the crystal structure of the MPP-presequence complex, a plausible model of preEGFP associated with MPP was constructed in silico. The modelled structure indicated that amino acid residues on the C-terminal side of the cleavage site in the preprotein were orientated tail out from the large cavity of MPP and interacted with the glycine-rich loop of α-MPP. Thus, MPP orientates preproteins at the specific cleft between the catalytic domain and the flexible glycine-rich loop which seems to pinch the extended polypeptide.

本文言語英語
ページ(範囲)889-895
ページ数7
ジャーナルJournal of biochemistry
141
6
DOI
出版ステータス出版済み - 6 2007

All Science Journal Classification (ASJC) codes

  • 生化学
  • 分子生物学

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