Self-assembly of large, ordered lamellae from non-bilayer lipids and integral membrane proteinsin vitro

Abstract

In many biological membranes, the major lipids are ''non-bilayer lipids,'' which in purified form cannot be arranged in a lamellar structure. The structural and functional roles of these lipids are poorly understood. This work demonstrates that the in vitro association of the two main components of a membrane, the non-bilayer lipid monogalactosyldiacylglycerol (MGDG) and the chlorophyll-a͞b light-harvesting antenna protein of photosystem II (LHCII) of pea thylakoids, leads to the formation of large, ordered lamellar structures: (i) thin-section electron microscopy and circular dichroism spectroscopy reveal that the addition of MGDG induces the transformation of isolated, disordered macroaggregates of LHCII into stacked lamellar aggregates with a long-range chiral order of the complexes; (ii) small-angle x-ray scattering discloses that LHCII perturbs the structure of the pure lipid and destroys the inverted hexagonal phase; and (iii) an analysis of electron micrographs of negatively stained 2D crystals indicates that in MGDG-LHCII the complexes are found in an ordered macroarray. It is proposed that, by limiting the space available for MGDG in the macroaggregate, LHCII inhibits formation of the inverted hexagonal phase of lipids; in thylakoids, a spatial limitation is likely to be imposed by the high concentration of membrane-associated proteins.circular dichroism ͉ chloroplast thylakoid membranes ͉ electron microscopy ͉ light-harvesting complex ͉ lipid-protein interactions T he lamellar organization of biological membranes provides a structural matrix for various proteins and controls the permeability of organic molecules, water, and ions; it also prevents nonspecific protein-protein aggregation, whereas it allows protein diffusion and conformational changes in the membrane. However, biomembranes usually contain substantial amounts of non-bilayer lipids, which in purified form assume nonlamellar structures. In fact, in many membranes, e.g., thylakoid membranes of chloroplasts, membranes of Escherichia coli, rhodopsin, and mitochondria, non-bilayer lipids constitute about half or more of the total lipid content. It is well established that the physical and functional properties of these membranes depend to a large extent on protein-lipid interactions (1, 2). There are a few examples showing that lipid polymorphism can be modulated by proteins, and, in some cases, small unilamellar vesicles can be reconstituted from non-bilayer lipids and membrane proteins (e.g., refs. 3 and 4). However, the structural role of large amounts of non-bilayer lipids has remained enigmatic, and the assembly of extended bilayer lamellae from proteins and predominantly non-bilayer lipids is poorly understood (1, 2, 5). In this work, we use a simple system, the two main constituents of pea thylakoid membranes, purified non-bilayer lipids and isolated protein complexes, to demonstrate that the formation of a large, ordered lamellar structure is possible even in the presence of large amounts of lipids.In chloroplast thylakoid membranes of g...

Molecular Architecture of Plant Thylakoids under Physiological and Light Stress Conditions: A Study of Lipid-Light-Harvesting Complex II Model Membranes

“…Most of these data suggest that the primary site of light-induced reorganizations is on the outer loop segments of LHCII, affecting the phosphorylation (Zer et al, 1999), the cation binding sites (Cseh et al, 2000;Garab and Mustardy, 2000), and the ability of stacking and lateral reorganizations (Dobrikova et al, 2003;Gruszecki et al, 2009b). Despite the differences in xanthophyll composition and phosphorylation status of the LHCII and LHCII-HL samples, both the fluorescence emission spectra and the fluorescence lifetimes are almost identical (see Supplemental Figure 8A online).…”

“…In the case of spinach (Spinacia oleracea), MGDG constitutes 57% of the thylakoid membrane lipids and DGDG constitutes 27% (Douce and Joyard, 1996). The formation of the relatively large areas of the planar lipid bilayers of the thylakoid membranes, such as in the case of the grana structures, is thought to be possible due to the relatively high fraction of transbilayer protein components, such as LHCII, which stabilize the bilayer structure Simidjiev et al, 2000).…”

The Arabidopsis IspH Homolog Is Involved in the Plastid Nonmevalonate Pathway of Isoprenoid Biosynthesis

“…6, D-I). The integration of the LHCs, which are mainly composed of chlorophylls, carotenoids, and apoproteins, is important for the development of thylakoids and the formation of grana stacking (Bartley and Scolnik, 1995;Von Wettstein et al, 1995;Simidjiev et al, 2000). Thus, the development of the chloroplast in the IspH-silencing cells may be arrested at various stages depending on the levels of silencing.…”

A Suppressor of fab1 Challenges Hypotheses on the Role of Thylakoid Unsaturation in Photosynthetic Function

“…Thus, one possibility is that changes in the shape of MGD molecules compensate for disruptive changes in the shape of PG molecules induced by the fab1 mutation, for example by altering the packing relationships between the thylakoid lipids and membrane proteins of the photosynthetic complexes (Gounaris and Barber, 1983;Simidjiev et al, 2000). Changes in the shape of lipid molecules is known to be important for a number of membrane functions including protein trafficking and membrane fusion (Rietveld et al, 1995;Chanturiya et al, 1997;Bruce, 1998).…”