title: A minimal peptide sequence that targets fluorescent and functional proteins into the mitochondrial intermembrane space.
abstract: Protein-based fluorescent and functional probes are widely used for real-time visualization, purification, and regulation of a variety of biological molecules. The protein-based probes can generally be targeted into subcellular compartments of eukaryotic cells by a particular short peptide sequence. Little is known, however, about the sequence that targets probes into the mitochondrial intermembrane space (IMS). To identify the IMS-targeting sequence, we developed a simple genetic screening method to discriminate the proteins localized in the IMS from those in the mitochondrial matrix, thereby revealing the minimum requisite sequence for the IMS targeting. An IMS-localized protein, Smac/DIABLO, was randomly mutated, and the mitochondrial localization of each mutant was analyzed. We found that the four residues of Ala-Val-Pro-Ile are required for IMS localization, and a sequence of these four residues fused with matrix-targeting signals is sufficient for targeting the Smac/DIABLO into the IMS. The sequence was shown to readily direct three dissimilar proteins of interest to the IMS, which will open avenues to elucidating the functions of the IMS in live cells.
authors.affiliation: Department of Chemistry, School of Science, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan.
authors.forename: Takeaki
authors.initials: T
authors.lastname: Ozawa
authors.forename: Yutaka
authors.initials: Y
authors.lastname: Natori
authors.forename: Yusuke
authors.initials: Y
authors.lastname: Sako
authors.forename: Haruko
authors.initials: H
authors.lastname: Kuroiwa
authors.forename: Tsuneyoshi
authors.initials: T
authors.lastname: Kuroiwa
authors.forename: Yoshio
authors.initials: Y
authors.lastname: Umezawa
mesh_terms: D000595:Amino Acid Sequence; D051017:Apoptosis Regulatory Proteins; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D008164:Luminescent Proteins / Q000378:metabolism*; D051336:Mitochondrial Membranes / Q000378:metabolism*; D024101:Mitochondrial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D016296:Mutagenesis; D010446:Peptide Fragments / Q000378:metabolism; D021381:Protein Transport
chemical_list: D051017:Apoptosis Regulatory Proteins; C412403:DIABLO protein, human; D047908:Intracellular Signaling Peptides and Proteins; D008164:Luminescent Proteins; D024101:Mitochondrial Proteins; D010446:Peptide Fragments