The specificity of the shift was confirmed using antibodies against the nuclear protein ENP1 (Figure?7E; ENP1), which is not related to protein translocation into chloroplasts (Missbach et al

The specificity of the shift was confirmed using antibodies against the nuclear protein ENP1 (Figure?7E; ENP1), which is not related to protein translocation into chloroplasts (Missbach et al., 2013). by mutation of the last -strand, suggesting that this strand contributes to the insertion signal. These findings shed light on the elements and complexes involved in plastidic -barrel protein import. Introduction Most organellar proteins are synthesized at cytosolic ribosomes as precursor proteins. Consequently, the multiplicity of possible target destinations demands a high targeting accuracy. This specificity is generally guaranteed by the presence of a targeting signal that ensures the correct targeting and insertion of the proteins into the respective organelle and suborganellar compartments (Schatz and Dobberstein, 1996). This specificity appears to be high, as even plastids and mitochondria, which share many enzymatic actions due to their common Gram-negative bacterial origin, share only a small number (-)-DHMEQ of dual-targeted proteins (Carrie et al., 2009). Most precursor proteins of the mitochondrial matrix and the chloroplast stroma contain an N-terminal topogenic signal called the presequence or transit peptide, respectively, that is cleaved after successful translocation by intraorganellar processing peptidases (Schleiff and Becker, 2011; Jarvis and Lpez-Juez, 2013). These topogenic signals have an overall positive charge and the tendency to form an amphiphilic -helix (Bruce, 2001; Chacinska et al., 2009). In addition, chloroplast transit (-)-DHMEQ peptides are enriched in hydroxylated amino acids, which can be phosphorylated by cytosolic kinases (Martin et al., 2006). Presequences and transit peptides are divergent in length and primary structure, leading to the hypothesis that instead of a specific-sequence motif, a structural element or physiochemical pattern is (-)-DHMEQ important for recognition around the organellar receptors (Schleiff and Becker, 2011; Kunze and Berger, 2015). Open in a separate windows The topogenic (-)-DHMEQ signal of some proteins, especially outer membrane proteins, appears to be distinct from this general scheme. One class with such distinct signals consists of the -barrel proteins in the outer membranes of mitochondria and chloroplasts. In mitochondria of the yeast that are annotated according to their chromosomal locations: (Jackson-Constan and Keegstra, 2001). is usually a pseudogene and is thought to be expressed in specific organs or only under specific conditions (Baldwin et al., 2005). TOC75-III constitutes the pore-forming translocation channel of the translocon of the outer membrane of chloroplasts (TOC; Schleiff and Becker, 2011), while TOC75-V (Eckart et al., 2002; Patel et al., 2008) is usually thought to function in -barrel protein insertion (Schleiff and Soll, 2005), which, however, still needs to be confirmed. As resulted in targeting of this chimeric protein to mitochondria in protoplasts (Jores et al., 2016). With respect to the translocation of plastidic -barrel proteins, recent findings revealed a Mouse monoclonal to CD95(FITC) possible intermediate in the IMS (Klinger et al., 2019) derived by TOC-dependent translocation across the outer membrane (Day et al., 2019). All subsequent actions are currently (-)-DHMEQ unclear. To gain insights into the targeting and insertion of -barrel proteins in plants, we utilized a protoplast-based and an import assay to analyze the translocation of plastidic -barrel proteins using OEP21, OEP24, OEP37, and P39 from pea (and (Machettira et al., 2011a, Ulrich et al., 2012; Jores et al., 2016; Klinger et al., 2019). Recent topology prediction suggested that mesophyll protoplasts or onion (protoplasts and epidermal cells. A, mesophyll protoplasts were co-transformed with.