Pore structure of channel proteins

Four-Helix Bundles A Plausible Model for the Pore Structure of Channel Proteins... 

For all other members of the TRP family it still has to be shown whether the presumptive pore loop or other protein domains actually line the ion conducting pathway of the channel. Based on the results showing that expression of most TRPC channels yields currents carried by Na+and Ca2+, and that expression of TRPM4 and TRPM7 channels yields currents carried by Na+ but not by Ca2+(TRPM4) or even currents carried by Mg2+or Ca2+(TRPM7), it seems likely that the pore structures of these channel proteins vary considerably. 

Fig. 16.13. Pore structure of the acetylcholine receptor, based on electron microscopy studies. a) Electron density map of the acetylcholine receptor of the postsynaptic membrane of the electric organ of the ray Torpedo californicus, based on electron microscopy studies. The receptor has a long funnel-like structure in the extracellular region, which narrows at the center of the pore. A smaller funnel structure is observed in the cytoplasmic region of the receptor. Another protein is situated on the cytoplasmic side. The long arrow indicates the direction of ion passage and the small arrow shows the postulated binding site for acetylcholine, b) Schematic representation of the acetylcholine receptor with the M2 hehx as the central block in the ion channel. According to Unwin, (1993).

The ATP-gated ion channels are a family of channel proteins. ATP-gated ion channels form from three separate proteins, each of which contains two transmembrane a-helices. The collective total of six helices form a pore through the membrane and allow the passage of ions. ATP-gated ion channels therefore involve quaternary structure based on the relative orientation of the three protein subunits.3... 

Our knowledge about the composition and structure of membrane proteins comes from isolation and purification attempts as well as from experiments with model compounds (e.g., ionophores), all of which indicate that we should expect substantial diversity. In terms of size, we can anticipate a range from perhaps 1000 to several 100,000 daltons. We may also assume that most membrane proteins are amphipathic. The hydrophobic region could be provided by a ten-unit sequence of non-polar (CAPALDI 6e VANDERKOOI, 1972) amino acids or by a large hydrocarbon chain as in the case of the proteolipids. It is often assumed that the hydrophobic regions of the membrane proteins are immersed in the lipid bi-layer while the more polar moieties face the cytoplasmic compartments. This assumption, however, is backed insufficiently by experimentation, because proteins may form hydrophilic channels (pores) across the membrane which are lined inside with hydrophilic charged groups. 

Plate 29 Structure of membrane proteins. Membrane peptides often self assemble into controlled oligomerie forms to make molecular selective channels whose structure can be very difficult to resolve at the molecular level by most methods, although solid state NMR methods can make a contribution to their functional and structural description. Here the pentameric funnel-Hke bundle of the M2-peptide from the nicotinic acedy choline receptor has been resolved from NMR studies of oriented M2 peptides in lipid bilayers. The funnel has a wide mouth at the N-terminal, intracellular side of the pore. The pore lining residues has also been modelled and distances between residues in the channel estimated. The a-carbon backbone is in cyan, acidic residues in red and basic residues in blue, polar residues in yellow and lipophilic residues in purple. (Figure adapted from Opella et al., (1999) Nature St. Biology, 6 374-379). See Membranes Studied by NMR Spectroscopy. 

Whereas the main challenge for the first bilayer simulations has been to obtain stable bilayers with properties (e.g., densities) which compare well with experiments, more and more complex problems can be tackled nowadays. For example, lipid bilayers were set up and compared in different phases (the fluid, the gel, the ripple phase) [67,68,76,81]. The formation of large pores and the structure of water in these water channels have been studied [80,81], and the forces acting on lipids which are pulled out of a membrane have been measured [82]. The bilayer systems themselves are also becoming more complex. Bilayers made of complicated amphiphiles such as unsaturated lipids have been considered [83,84]. The effect of adding cholesterol has been investigated [85,86]. An increasing number of studies are concerned with the important complex of hpid/protein interactions [87-89] and, in particular, with the structure of ion channels [90-92]. 

Concerning the nature and structure of such amyloid peptide or protein channels, oligomers with annular morphologies have in fact been observed by EM for a-synuclein (Lashuel et al., 2002) and equine lysozyme (Malisauskas et al., 2003) even in the absence of any lipids or membranes. Channel-like structures have also been reconstituted in liposomes and observed by SFM for A/ i 4o, A/ j 42, human amylin, a-synuclein, ABri, ADan, and serum amyloid A (Fig. 5A Lin et al., 2001 Quist et al., 2005). Doughnut-shaped structures with a diameter of 10-12 nm and a central hole size of 1-2 nm (Fig. 5B) were imaged on top of lipid membranes (Quist et al., 2005). However, the radius of curvature of the SFM tips meant that it is not possible to say whether the pores were really traversing the lipid bilayer. 

Lashuel, H. A., Petre, B. M., Wall, J., Simon, M., Nowak, R. J., Walz, T., and Lansbury, P. T., Jr. (2002). Alpha-synuclein, especially the Parkinson s disease-associated mutants, forms pore-like annular and tubular protofibrils./. Mol. Biol. 322,1089-1102. LeVine, H. (1993). Thioflavine T interaction with synthetic Alzheimer s disease beta-amyloid peptides Detection of amyloid aggregation in solution. Protein Sci. 2, 404—410. Lin, H., Bhatia, R., and Lai, R. (2001). Amyloid beta protein forms ion channels Implications for Alzheimer s disease pathophysiology. FASEB J. 15, 2433-2444. Lorenzo, A., and Yankner, B. A. (1994). Beta-amyloid neurotoxicity requires fibril formation and is inhibited by Congo red. Proc. Natl. Acad. Sci. USA 91, 12243-12247. Luhrs, T., Ritter, C., Adrian, M., Riek-Loher, D., Bohrmann, B., Dobeli, H., Schubert, D., and Riek, R. (2005). 3D structure of Alzheimer s amyl o id-( be la) (1—12) fibrils. Proc. Natl. Acad. Sci. USA 102, 17342-17347. 

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