Thylakoid membrane protein insertion

Chloroplasts in higher plants have three membranes the outer and inner envelope membranes and the thylakoid membrane. Very little is known about membrane protein assembly into the two envelope membranes (Soil and Tien, 1998). The thylakoid has been better studied and in fact appears to use mechanisms very similar to those found in E. coli for membrane protein insertion (Dalbey and Robinson, 1999). Thus, SRP, SecA, SecYEG, YidC, and Tat homologues are all present in the thylakoid membrane or in the stroma (the Tat system was first identified in thylakoids, in fact). In contrast to E. coli, however, there are thylakoid proteins that appear to insert spontaneously into the membrane, insofar as no requirement for any of the known translocation machineries has been detected (Mant et al, 2001). 

Dalbey, R. E., and A. Kuhn. 2000. Evolutionarily related insertion pathways of bacterial, mitochondrial, and thylakoid membrane proteins. Ann. Rev. Cell Devel. Biol. 16 51-87. 

The transport of proteins into chloroplasts also occurs by more than one mechanism. An SRP-dependent pathway may be needed only for insertion of proteins into membranes.594 Other proteins, among which are the 23-kDa and 16-kDa photosystem II proteins (Chapter 23), enter by a pathway related to the Tat pathway of bacteria. In thylakoids this pathway is directly dependent upon the large pH difference (A pH) across die thylakoid membrane. In contrast to the bacterial Sec pathway, the A pH pathway seems to be able to transport completely folded proteins. 

Some proteins enter membranes immediately after synthesis. The translocon channel is not required. However, in E. coli an additional protein YidC is needed.603 Homologs of this protein are found in mitochondria (Oxal protein) and in thylakoid membranes of chloroplasts (Alb3 protein).608 These proteins may function in cotranslational insertion. If a protein carries a... 

Lysed chloroplasts from norflurazon-treated pea plants, containing only 5% oftheirnormal carotenoid contents, stably inserted less than 10% of Lhcbl into their thylakoid membranes compared to control experiments with non-treated plants (Dahlin and timko, 1994). This indicates either that the low amounts of carotenoids (and/or Chi b) in these chloroplasts were insufficient to stabilize newly inserted Lhcbl, or that the lack of pigments inhibited the insertion of Lhcbl into the thylakoid. Carotenoid deficiency due to norfluorazon treatment appears also to impair protein import into isolated chloroplasts. 

An involvement of carotenoids in the thylakoid insertion of light-harvesting proteins has also been concluded from a pulse-chase measurement of the rate of protein insertion into membranes that have reduced amounts of carotenoids, due to norflurazon treatment (Dahlin and Timko, 1997). It should be kept in mind that carotenoids may also control other biosynthetic steps in the synthesis and assembly of light-harvesting complexes such as the import of the protein precursors into plastids (Dahlin, 1993 Dahlin and Franz6n, 1997). 

Salih G, Wiklund R, Tyystjarvi T et al. Constructed deletions in lumen-exposed regions of the D1 protein in the cyanobacterium Synechocystis 6803 Effects on 01 insertion and accumulation in the thylakoid membranes, and on Photosystem II assembly. Photosynth Res 1996 49 131-140. 

One important aspect of LHCII that specifically relates to other aspects discussed in the present review is the question of how the membrane environment (lipid composition, membrane curvature, etc.) affects the association of LHCII monomers to form trimers and the assembly of these trimers into the antenna complex around the photosynthetic reaction centers. The nonbilayer-forming lipid monogalactosyldiacylglycerol (MGDG) constitutes half of the thylakoid membrane. This membrane maintains its lamellar structure only with proteins inserted, predominantly LHCII which, due to its concave shape, eases the curvature pressure exerted by MGDG. It has been suggested that this curvature pressure is a driving force for protein interaction in the membrane [317] however, because it is not known whether, e.g., the formation of supercomplexes of LHCII trimers eases or increases curvature pressure, it is unclear whether MGDG (or other curvature... 

Rieske protain/centar an iron-sulphur protein first isolat from Complex III of the mitochondrial electron transport chain, in which it occurs with cytochromes b and C) [J.S, Rieske el al. Biochem. Biophys Res Commun. IS (1%4) 338-344], but which has now been found in the equivalent cytochrome be complexes in the bacterial plasma membrane and the chloroplast thylakoid membrane. The latter, known as the cytochrome bff complex, partidpates in cyclic and noncyclic electron flow in the light phase of photosynthesis (see Photosynthesis). All Rieske proteins are one-electron redox systems with a standanl redox potential in the + 0.2 to + 0.3V range and have a (2Fe-2S] center, a single membrane-spanning a-helix, and a characteristic electron spin resonance (ESR) spectrum. The chloroplastidic R.p/c, with a M, of - 20,000, is smaller than that of the mitochondnon. It is encoded in the nucleus, synthesized in the cytoplasm and translocated to the chloroplast, where it is inserted into the thylakoid membrane. Within the thylakoid membrane its [2Fe-2S] redox centre (near to its C-terminus) can readily pass electrons to cytochrome /, a c-type cytochrome that projects from the luminal surface cytochrome / then passes electrons to plastocyanin (see) dissolved in the aqueous milieu of the thylakoid lumen. 

The transmembrane asymmetry of acyl lipids raises several interesting questions. The most intriguing ones concern the origin of this asymmetry and the reciprocal influence of proteins and acyl lipids on their respective organization. Another way to put it is to ask whether the insertion of new proteins in the thylakoid membrane induces a new or maintains the same acyl lipid asymmetry. The knowledge of the distribution of acyl lipids in prothylakoids may provide answers to the above questions [9,10]. Frothylakoids are the precursors of mature thylakoids but their function and protein composition are quite different, e.g. prothylakoids are devoid of... 

FIGURE 1. Insertion of LHCP into the thylakoids in the correct orientation following transport into the chloroplast by the transit peptide pS. RNA was transcribed from genes coding for pLHCP and the hybrid protein was translated in a wheat germ lysate, and 5ie products (lanes 1, panel A, pLHCP panel B, hybrid) were incubated with chloroplasts in an import reaction, llie total membrane-associated proteins are shown before (lanes 2) and after (lanes 3) trypsin treatment. 

Figure 3. Insertion of labeled ol-HCP into isolated thylakoids was carried out for 30 minutes at luC. The lab ed precursor was then chased with excess of unlabeled pLHCP at 2. Fractions of 700 ul were removed and fractionated into grana (G)- and stroma (S)-ldmellae immediatly after application of unlabeled precursor, zero time, or after 10, 20, and 40 minutes. Equal amounts of the two membranal fractions (10 ug Chi) were analyzed by SDS-PA6E followed by protein staining (a) and fluorography (b).

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