7), but not Sec pathway substrates or LHCP (data not shown)

7), but not Sec pathway substrates or LHCP (data not shown). in 10-collapse excess over active translocation sites. Antibodies to either Tha4 or Hcf106 inhibited translocation of four known Delta pH pathway substrate proteins, but not of Sec pathway or SRP pathway substrates. This suggests that Tha4 and Hcf106 operate either in series or as subunits of a heteromultimeric complex. cpSecY antibodies inhibited translocation of Sec pathway substrates but not of Delta pH or SRP pathway substrates. These studies provide the 1st biochemical evidence that Tha4 and Hcf106 are specific components of the Hydroxocobalamin (Vitamin B12a) Delta pH pathway and provide one line of evidence that cpSecY is used specifically from the Sec pathway. Keywords:chloroplast protein transport, twin arginine, SecY, Hcf106, Tha4 Proteintranslocation across and into membranes is definitely a fundamental cellular process, responsible for localization of 50% of the proteins inside a eukaryotic cell.Schatz and Dobberstein 1996classified eukaryotic protein translocation into import-type systems and export-type systems. Import systems translocate proteins from your cytosol into organelles and appear to have developed after endosymbiosis. Export systems are derived from prokaryotic systems for exporting proteins across the cytoplasmic membrane and are present in the ER, the mitochondrial inner membrane, and chloroplast thylakoid membranes. Some proteins such as nucleus-encoded thylakoid proteins are localized by sequential action of an import system and an export system. Recently, it has been shown that there are several different export systems present in thylakoid membranes andEscherichia coliplasma membranes (for evaluations seeSettles and Martienssen 1998;Dalbey and Robinson 1999;Keegstra and Cline 1999). In the case of thylakoids, these systems (or pathways) translocate unique subgroups of proteins and may be distinguished by energy and stromal protein requirements, by competition with overexpressed precursors, and by recognized components of the translocation machinery (Keegstra and Cline 1999). One pathway, termed the thylakoid Sec pathway, is responsible for translocation of plastocyanin (Personal computer),1OE33, PSI-F, and the plastid-encoded cytochrome F. The thylakoid Sec pathway appears to be homologous to the well-studied bacterial Sec pathway (Economou 1998) because it requires ATP and cpSecA, a homologue of the bacterial SecA protein, and is stimulated from the thylakoidal pH gradient (Keegstra and Cline 1999). The Sec pathway also utilizes membrane-bound machinery (Robinson et al. 1996), which is assumed to consist of cpSecY and cpSecE, homologues of the bacterial translocon proteins, SecY and Hydroxocobalamin (Vitamin B12a) SecE. A second pathway, the chloroplast transmission acknowledgement particle (SRP) Tnfsf10 pathway, is responsible for targeting a family of integral thylakoid proteins, the LHCPs (Li et al. 1995), and probably also the plastid-encoded D1 protein (Nilsson et al. 1999). The SRP pathway appears to be homologous to the bacterial and ER SRP pathways because it utilizes a chloroplast homologue of the SRP54 protein, forms a soluble complex with LHCP substrates in the stroma, and requires GTP for integration (Keegstra and Cline 1999). The chloroplast SRP system possesses some unique features. It can operate posttranslationally, it lacks an connected RNA, and it possesses a novel protein component, cpSRP43, which apparently confers cpSRP54 with its posttranslational ability (Schuenemann et al. 1998). The chloroplast SRP system utilizes membrane protein(s) (Robinson et al. 1996) that have not Hydroxocobalamin (Vitamin B12a) yet been recognized. One possibility is that the chloroplast SRP docks having a Sec-like translocon, similar to the ER SRP (Walter and Johnson 1994) and the bacterial SRP (Valent et al. 1998). A third thylakoid pathway is definitely termed the Delta pH pathway because it utilizes the thylakoidal pH gradient as only energy source for transport of lumenal proteins (Cline et al. 1992). The Delta pH pathway is responsible for transport of OE23, OE17, PSI-N, and PSII-T and exhibits several special and unusual features. It requires neither NTPs nor soluble protein factors. Its substrates possess conserved twin arginines in their transmission peptide that, where examined, are essential for transport (Chaddock et al. 1995;Henry et al. 1997). Also, it seems capable of translocating tightly folded proteins (Clark and Theg Hydroxocobalamin (Vitamin B12a) 1997;Hynds et al. 1998). The Delta pH system was initially thought.

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