370 During the past year, research on helical membrane proteins has brought insights into the use of deviations from canonical α-helical conformation to support function and the further investigation of the sequestration of protein regions from the lipid bilayer to enhance these structural alternatives. Also, the structural roles of polar sidechains, the identification of motifs in helix interactions and the significance of certain topologies on a genome-wide scale have been further explored. Addresses Department of Molecular Biophysics and Biochemistry, Yale University, 266 Whitney Avenue, PO Box 208114, New Haven, CT 06520-8114, USA *e-mail: don@paradigm.csb.yale.edu Current Opinion in Structural Biology 2001, 11:370—376 0959-440X/01/$ —see front matter © 2001 Elsevier Science Ltd. All rights reserved. Abbreviations GpA glycophorin A GPCR G-protein-coupled receptor PDB Protein Data Bank Introduction A membrane protein is exposed to a heterogeneous envi- ronment, where the transmembrane regions are embedded in a phospholipid bilayer and the extramembrane domains are surrounded by water. The water-exposed polypeptide can adopt a diverse array of folds, whereas the physical and chemical constraints imposed by the lipid bilayer appear to restrict the structural diversity of the embedded protein domain (for recent reviews on membrane protein structure and folding, see [1–5]). All membrane protein structures solved to date show that transmembrane domains fold as either single α helices, bundles of α helices or β strands assembled in β barrels (Figure 1). In this review, we will focus on α-helical membrane proteins. The folding of α-helical membrane proteins has been con- ceptualized as a two-stage process. Initially, hydrophobic polypeptide segments form independently stable trans- membrane α helices across the membrane and can be regarded as domains. Subsequently, these helical domains assemble laterally to form the native protein [1,3]. Underlying this model is the postulate that one possible approach towards understanding helical membrane protein structure is to analyze the structure of individual transmem- brane helices and, subsequently, the interactions and motifs that drive their association. Here, we review emerging themes regarding the structure of transmembrane α helices and their interactions. As revealed by the membrane protein structures published in the past year, deviations from the canonical α helix are relatively common and occur mostly to fulfill certain important functional roles. Furthermore, the relevance of polar interactions between transmembrane helices is becoming clearer from structural as well as bio- physical evidence. We also explore the transmembrane helix–helix packing motifs that have recently become more evident. Finally, we discuss the occurrence of predominant helical membrane protein folds, as revealed by genome-wide analysis of membrane proteins in the past year. Deviations from the ‘canonical’ structure of transmembrane α helices have functional consequences Current membrane protein topology prediction methods achieve over 75% accuracy in predicting the full topologies of α-helical membrane proteins [6,7]. Using these methods, in combination with experimental approaches, a large data- base of predicted transmembrane domains has been generated. Statistical analysis of these databases reveals that, on average, transmembrane helices are hydrophobic and composed of approximately 20–30 amino acids. Their central region is rich in aliphatic residues and phenylala- nines, with short border regions enriched in tryptophan and tyrosine [8,9]. This view of the composition of a typical transmembrane helix was reinforced when recent three- dimensional structures of membrane proteins were analyzed [10,11]. From a structural perspective, the elucidated struc- tures reveal that, in a typical transmembrane helix, hydrogen bonds are intrahelical, with interactions between residue i and residue i + 4. However, deviations in helix con- formation, length and composition do occur. Recently determined structures provide interesting examples of some of these deviations. A π-bulge, which is a deformation from a regular α-helical conformation, arises when backbone hydrogen bonds occur between residue i and residue i + 5 (Figure 2). In the recently determined structure of the light-driven anion pump halorhodopsin, helices E and G are distorted by π-bulges [12]. The high-resolution structure of the homol- ogous light-driven proton pump bacteriorhodopsin also reveals a π-bulge in helix G [13 • ]. Another interesting type of deformation is helix unwinding, examples of which can be observed in two transmembrane helices (M4 and M6) of the calcium pump of the sarcoplasmic reticulum [14 • ]. Finally, proline-induced kinks are often observed in struc- tures, examples of which are found in helices E and C of bacteriorhodopsin [13 • ], in helix 6 of the G-protein- coupled receptor (GPCR) rhodopsin [15 •• ] and in one of the helices of the fumarate reductase from bacteria [16,17]. Structurally speaking, these distortions in α-helicity have at least two consequences: local conformational instability Helical membrane proteins: diversity of functions in the context of simple architecture Iban Ubarretxena-Belandia and Donald M Engelman*