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Targeting Capacity and Conservation of PreP Homologues Localization in Mitochondria of Different Species
Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
Visa övriga samt affilieringar
2011 (Engelska)Ingår i: Journal of Molecular Biology, ISSN 0022-2836, E-ISSN 1089-8638, Vol. 410, nr 3, s. 400-410Artikel i tidskrift (Refereegranskat) Published
Abstract [en]

Mitochondrial presequences and other unstructured peptides are degraded inside mitochondria by presequence proteases (PrePs) identified in Arabidopsis thaliana (AtPreP), humans (hPreP), and yeast (Cym1/Mop112). The presequences of A. thaliana and human PreP are predicted to consist of 85 and 29 amino acids, respectively, whereas the Saccharomyces cerevisiae Cym1/Mop112 presequence contains only 7 residues. These differences may explain the reported targeting of homologous proteins to different mitochondrial subcompartments. Here we have investigated the targeting capacity of the PreP homologues' presequences. We have produced fusion constructs containing N-terminal portions of AtPreP(1-125), hPreP(1-69), and Cym1(1-40) coupled to green fluorescent protein (GFP) and studied their import into isolated plant, mammalian, and yeast mitochondria, followed by mitochondrial subfractionation. Whereas the AtPreP presequence has the capacity to target GFP into the mitochondrial matrix of all three species, the hPreP presequence only targets GFP to the matrix of mammalian and yeast mitochondria. The Cym1/Mop112 presequence has an overall much weaker targeting capacity and only ensures mitochondrial sorting in its host species yeast. Revisiting the submitochondrial localization of Cym1 revealed that endogenous Cym1/Mop112 is localized to the matrix space, as has been previously reported for the plant and human homologues. Moreover, complementation studies in yeast show that native AtPreP restores the growth phenotype of yeast cells lacking Cym1, demonstrating functional conservation.

Ort, förlag, år, upplaga, sidor
2011. Vol. 410, nr 3, s. 400-410
Nyckelord [en]
mitochondrial import, presequence, PreP, Cym1/Mop112, peptide degradation
Nationell ämneskategori
Biokemi och molekylärbiologi Biofysik
Identifikatorer
URN: urn:nbn:se:su:diva-66595DOI: 10.1016/j.jmb.2011.05.009ISI: 000292718400003OAI: oai:DiVA.org:su-66595DiVA, id: diva2:468250
Anmärkning
authorCount :9Tillgänglig från: 2011-12-20 Skapad: 2011-12-20 Senast uppdaterad: 2017-12-08Bibliografiskt granskad
Ingår i avhandling
1. Dual targeting of proteins to mitochondria and chloroplasts: Characterization of dual targeting peptides and their interaction with organellar receptors
Öppna denna publikation i ny flik eller fönster >>Dual targeting of proteins to mitochondria and chloroplasts: Characterization of dual targeting peptides and their interaction with organellar receptors
2014 (Engelska)Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
Abstract [en]

Most mitochondrial and chloroplastic proteins are synthesized in the cytosol as precursor proteins with an N-terminal targeting peptide (TP), which directs them to the correct organelle. There is also a group of proteins that are dual targeted to mitochondria and chloroplasts using an ambiguous N-terminal dual targeting peptide (dTP). The aim of this thesis was to characterize dTPs with respect to physicochemical features, sequence patterns, structural properties and interaction with the mitochondrial and chloroplastic receptors.

We have used different statistical methods, including a multivariate data analysis (MVDA) to analyse all available dTPs and compare them to organelle-specific TPs of proteome-identified mitochondrial and chloroplastic proteins from Arabidopsis thaliana. The overall amino acid sequence patterns of dTPs were intermediate between mitochondrial targeting peptides (mTPs) and chloroplastic targeting peptides (cTPs) but the greatest differences in amino acid composition were found within the very N-terminal region of dTPs, where especially arginines are highly overrepresented in relation to cTPs. Interestingly, introducing arginines to the dTPs showed clustering towards the mTPs in silico and resulted in inhibition of chloroplast import in vitro, suggesting that positive charges in the N-terminal region of TPs may function as an 'avoidance signal' for chloroplast import.

Studies with the dTP of threonyl-tRNA synthetase (ThrRS-dTP) revealed that 60 amino acids were required to confer dual targeting. The purified ThrRS-dTP(2-60) inhibited import of organelle-specific proteins, providing evidence that dual and organelle-specific proteins use the same organellar import pathways. CD spectra indicated that ThrRS-dTP(2-60) has the propensity to form a-helical structure in membrane mimetic environments. Further, NMR investigations of interaction profiles of ThrRS-dTP(2-60) with the mitochondrial Tom20 and the chloroplastic Toc34 receptor demonstrated that the mode of the recognition of a dual targeting peptide by mitochondrial and chloroplastic receptors is different. Our studies provide thorough characterization of dTPs and present for the first time dTP-organellar receptor interactions on the molecular level.

Ort, förlag, år, upplaga, sidor
Stockholm: Department of Biochemistry and Biophysics, Stockholm University, 2014. s. 76
Nationell ämneskategori
Biokemi och molekylärbiologi
Forskningsämne
biokemi
Identifikatorer
urn:nbn:se:su:diva-108471 (URN)978-91-7649-027-3 (ISBN)
Disputation
2014-12-12, Magnélisalen, Kemiska övningslaboratoriet, Svante Arrhenius väg 16 B, Stockholm, 10:00 (Engelska)
Opponent
Handledare
Anmärkning

At the time of the doctoral defense, the following paper was unpublished and had a status as follows: Paper 5: Manuscript.

Tillgänglig från: 2014-11-20 Skapad: 2014-10-28 Senast uppdaterad: 2014-12-17Bibliografiskt granskad

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Alikhani, NyoshaBerglund, Anna-KarinSpånning, ErikaGlaser, Elzbieta
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Institutionen för biokemi och biofysik
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