Transmembrane elements

The transmembrane receptors span the ca. 5 mn thick phospholipid bilayer of the cell membrane with structural portions known as transmembrane elements. The inner of a phosphohpid layer is hydrophobic and, correspondingly, the surface of the structural elements that come into contact with the iimer of the phospholipid double layer also has hydrophobic character. 

The transmembrane domain may be made up of one or many transmembrane elements. Generally, the transmembrane elements include 20-25 mostly hydrophobic amino acids. At the interface with aqueous medium, we often find hydrophilic amino acids in contact with the polar head groups of the phospholipids. In addition, they mediate distinct fixing of the transmembrane section in the phospholipid double layer. A sequence of 20-25 hydrophobic amino acids is seen as characteristic for membrane-spaiming elements. This property is used in analysis of protein sequences, to predict possible transmembrane elements in so-called hydropathy plots". 

High resolution structural information about the transmembrane elements of membrane receptors is not currently available, since it is not yet possible to obtain transmembrane receptors in crystalline form for structural analysis. Due to the hydrophobic nature of the transmembrane elements, crystallization is very difficult. 

Thus, the important question of the secondary structure of the transmembrane elements can only be addressed with models and by structural comparison with other transmembrane proteins for which the structure has been resolved. Detailed information on the structure of transmembrane elements is available for the photoreaction center of Rhodopseudomonas viridis (review Deisenhofer and Michel, 1989), cytochrome c oxidase (Iwata et al., 1995) and the OmpF porin of E. coli (Cowan et al., 1992 Fig. 5.3), amongst others. In addition, high resolution electron microscopic investigations and X-ray studies of bacteriorhodopsin, a light-driven ion pump with seven transmembrane elements, have yielded valuable information on the structure and configuration of membrane-spaiming elements (Henderson et al., 1990 Kimura et al., 1997 Pebay-Peyrula et al., 1997 Fig. 5.4). With the successful crystallization of the photoreaction center of Rhodopseudomonas viridis, a membrane protein was displayed at atomic resolution for the first time (Deisenhofer et al., 1985). The membrane-... 

Fig. 5.3. Structure of the OmpF porin of E. coli. The porin is a bacterial membrane protein with P-sheet structures as transmembrane elements. The structure of a monomer of the OmpF porin is shown. In total, 16 P-bands are configured in the form of a cylinder and form the waUs of a pore through which selective passage of ions takes place. LI—L8 are long loops, Tl,2,3 and T7,8 are short bends (T turn) that fink the P-sheets. According to Cowan et al. (1992), with per-...

Hie transmembrane domain may consist of one or several transmembrane elements (see also Fig. 5.2). In the latter case, these are arranged in the form of bundles, as shown in Figure 5.4 for bacteriorhodopsin. In the case of ion channels, in which several subunits are involved in formation of the transmembrane domain (see acetylcholine receptor, Fig.16.12), prediction of the structure of the membrane portion is very difficult. The different transmembrane elements are no longer equivalent in these cases. Part of the element is involved in formation of the irmer wall of the pore, other structural elements form the surface to the hydrophobic irmer of the phospholipid bilayer. It is evident that the polarity requirements for the amino acid side chains vary according to the position of the transmembrane elements (see Chapter 16). 

In these cases, how the different transmembrane elements associate to an ordered structured transmembrane domain can only be discussed based on electron microscopy and crosslinking experiments and affinity marking. Predictions based on these experiments are really more like models. 

The occurrence of seven sequence segments, each made up of 20—25 amino acids, is characteristic for the G-protein coupled receptors. From this, it is assumed that they form transmembrane domains, and span the membrane in the form of a-helices. The transmembrane elements are linked by loops of various sizes on the outer and inner side. 

Figure 5.6 shows the primary sequence of the P-receptor for adrenaline with the assumed topology of the seven transmembrane hehces. The highest sequence homology of the G-protein coupled receptors is foimd in the transmembrane elements, whilst the hydrophilic loop regions show stronger divergence between different receptors. 

The subunit structure of the cytokine receptors is very variable. Amongst the cytokine receptors, there are receptors composed of one polypeptide and receptors made up of two or three different polypeptide chains (Fig. 11.3), which therefore have a hetero-oligomeric structure. The receptors have one transmembrane element per polypeptide chain. The NH2 terminus is located in the extracellular region whilst the CO OH terminus is intracellular. 

Fig. 12.4. Example of a two-component pathway in S. cerevisiae. Model of signal transdnction via the SLNl protein. The SLNl protein is a transmembrane protein with two transmembrane elements, which is assumed to exist as a dimer. The sensor domain and the regulator domain are localized on the same protein chain in the SLNl protein. The SLNl protein is activated by an extracellular signal (e.g., decrease in osmolarity). Autophosphorylation takes place on His (H) in the sensor domain and on Asp (D) in the regulator domain. A phosphate transfer takes place from the phosphohisti-dine to the effector protein SSKl. In the unphosphory-lated form, SSKl activates a MAPK pathway, which contains the protein kinase HOGl as a MAPK element. Various cellular reactions are triggered by HOGL If SSKl is phosphorylated in the course of activation of the two-component pathway, stimulation of the MAPK pathway is stopped. According to Swanson et al., (1994).

The charmel is a tetramer in which each subunit has two a-hehcal transmembrane elements. One helix of each subunit forms the irmer coating of the pore whilst the other helices form contacts to the phosphohpid bUayer, via hydrophobic residues. Hie helices are in the form of an. .inverted teepee with the broad opening oriented to the outside, into the extracellular region (Fig. 16.6). The loops between the helix pairs are on the extracellular side and are oriented inwards to the pore. These form the selectivity filter that discriminates between K and Na The narrow selectivity filter, which is only 12 A long, joins a large hydrophobic cavity and the inner pore, which is hydrophi-lically coated. 

Fig. 16.12. Subunit structure of the acetylcholine receptor, a) The acetylcholine receptor has the subunit structure 02 7 - The four transmembrane elements Ml—M4 are shown for the y subunit. The binding sites for acetylcholine (ACh) are located on the a-subunits. b) It is assumed that the inner wall of the ion pore is formed by M2 helices of the five subunits, c) Postulated configuration of the M2 helices in the narrowest region of the ion channel. In the closed state, five leucine residues (one per subunit) lie in the ion channel and hinder passage of ions. Above and below the block, there are negatively charged residues that serve as prefilters for ion passage.

High-resolution structural information about the transmembrane elements of transmembrane receptors could recently be obtained on the example of rhodopsin, the light-activated G protein coupled receptor of the vision process (Fig. 5.3). These data, together with earlier data on the structures of other transmembrane proteins (e.g., bacteriorhodopsin), have confirmed that a-helices are the principal structural building blocks of the transmembrane elements of membrane receptors. The transmembrane helices are composed of 20 - 30 hydrophobic amino acids with some polar... 

Fig. 6.3 Model ofthe domain structure ofthe natriuretic peptide receptor NPR, a receptor-type guanylyl cyclase. NPR is a dimeric transmembrane receptor which spans the membrane with two transmembrane elements. The extracytosolic domain comprises the ligand binding site and contains several disulfide bridges. The cytosolic part is composed of a kinase homology domain with multiple phosphorylation sites, an ATP binding site of unknown function and the catalytic guanylyl cyclase domain.

A total of seven tyrosine residues become phosphorylated during autophosphorylation. Two of these are located in the vicinity of the transmembrane element, three are in the activation segment of the catalytic domain, and a further two in the region of the C terminus. 

The cytokines of the TG F/> family are the activating ligands for the family of TGF/ receptors. This comprises two subfamilies, the type I (T/>R-I) and type II (T/>R-II) receptors. These are transmembrane proteins composed of an extracellular ligand-binding domain, a single transmembrane element, and a cytoplasmic Ser/Thr kinase do-... 

The protease responsible for the critical third proteolytic step is contained in a large protease complex named y-secretase (review Fortini, 2002). Proteins belonging to the presenilin family of proteases have been identified as the catalytic component of the y-secretase complex. The presenilins are transmembrane proteases with six transmembrane elements that use aspartate residues to cleave substrates. They have also been implicated in the processing of the Alzheimer precursor protein (APP), and mutations of the presenilins appear to contribute to the pathogenesis of Alzheimer s disease. In... 

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