Membrane protein structures: the known world expands

344

Membrane protein structures: the known world expands R Michael Garavito The structure determinations of the cytochrome bc 1 complex and the prokaryotic potassium channel demonstrate that a wider range of membrane proteins are now amenable to study by X-ray crystallography. Furthermore, the structures of porins and interfacial membrane proteins show that membrane structural biology is becoming a mature and productive field.

Figure 1

Addresses Department of Biochemistry, Michigan State University,East Lansing, MI 48824-1319, USA; [email protected]

Current Opinion in Bioteehnology 1998, 9:344-349 http://biomednet.com/elecref/0958166900900344 .c: Current Biology Publications ISSN 0958-1669 Abbreviations AQP aquaporin CL colipase PGHS prostaglandinH synthase PL

pancreatic Npase

Introduction "l\vo decades ago, thc gencral consensus was that intcgral m e m b r a n e proteins were not amenable to analysis bx, X-ray crystallography; howevcr, within two years, thc first reports of crystallizcd m c m b r a n c proteins appeared [1,2]. A decade ago, the structures of" only two m e m b r a n e protcins were known with any certainty: the bacterial photosynthetic rcaction ccnter I3,4] and bacteriorhodopsin, albeit at lower rcsolution [5]. Within the past ten years, e n o m m u s prngress has taken placc in the field of m e m b r a n e protein structural biology. T h e refinement of t h c techniques for membrane protein crvstallizatinn for X-ray diffraction [6,7] and advances in clectron microscopy of two-dimensional crystals [8"] has lcd to an cxplnsion in new structural informatinn abnut m e m b r a n e prntcins. This year has secn some imprcssivc ncx~ structures, important new insights into the mechanism of electron, inn and sohlte transport, and tcchnical advances for crystallizing m c m b r a n c proteins, which wc will discuss in this review. New from the electron transport chain: c y t o c h r o m e bc~ c o m p l e x With the recent reports of the structures of the bacterial [9] and beef heart [10,11] cytochromc c oxidases, the question was which clcctron transport complex would bc solvcd next. In mid-1997, Xia eta/. [12"] prcscnted the first description of the ubiqtiinnl-cytochrome ~ oxidoreductase or/~(1 complex from b e e f heart mitochondria. T h e hQ complex consists of 11 nonidentical subunits (2,165 amino acids): the crystal structure shows a pear-shaped symmetric dimer with a maximal width of 130 A and height of 155 ,~ (lgigure 1). T h e protein mass extends ()tit al)out 38 ,~ from the presumed transmembraile region on the cytochrnme (i sidc and 75 .~ tin the matrix side. T h c core 1 and 2 subunits, which arc

Current Opinion in Biotechnology I

A ribbon diagram of the cytochrome bc~ complex dimer as determined by Zhang et al. [13°']. The shaded area is the putative membranespanning domain; IMS and Matrix refer to the inner membrane space and matrix side of the inner mitochondrial membrane.

located on thc matrix side of the molcculc, displa,v homologous c(/[3 folds as well as intcrnal twoflild svmmetr\'. At the time of the report, the electron density maps had b c c n interpreted in terms of 1,900 amino acid residues, thrce home prosthetic groups, and one iron-sulfur center. T h e transmcmbrane region of the hr 1 complcx consists of 11 helical segments: 8 from cytochromc h. (mc from c v t o c h r o m e [1 , one from snbunit 7 and one frnm the ironsulfur protein. A recent report by Zhang rt a/. [13"1 also dcscribes the structures of the hr I complex from beef, rabbit, and chicken. T h e structural a g r e e m e n t bctxx.ecn these Dr i complexes and the complex reported by Xia eta/. [12"'1 is very good, ~iven some differences in the lncal qualit3 of clectron density. O n e major difference is that Zhang ri a/. [13"'] have a cnmplcte model for cytochromc q and subunit 8 and can resolve the Rieskc iron-sulfur protein xvcll e n o u g h to s t u d y its interactions with the rest of the complex. A comparativc analysis of the hc I complex in diffcrcnt states of oxidation has Icd to the novel Proposal that

MembraneproteinstructuresGaravito

domain m o v e m e n t accounts for some aspects of electron transfer [13"]. Xia #t a/. [ 1 2 - ] found that the inhibitor mvxothiazol lics in the ubiquinone Qo site between hcme bl. and the iron-sulfur center. Zhang eta/. [13"'] found that when a ligand occupying tire Qo site can make the appropriatc hydrogen bond, the Rieske protein is bound in the proximal (tight) position, near the (.)o site. \Vhcn this site is empty or is occupied by inhibitors, such as myxothiazol, the Rieskc protein is releascd to thc distal (loose) position near cytochromc q. Thus, Zhang eta/. propose that the catalvtic cycle requires a m o v e m e n t of the Ricske iron-sulfur cluster, first to the proximal position to take an electron from ubiqtlinol at the Qo site, then to the distal position to dcliver it to cvtochromc c 1. [lence, large-scale domain mo~cmcnts can play a role in electron transfer. ()nc interesting technical note is that both groups crystallizcd dctcrgent-solubilizcd hi! complexes usin R what could bc called 'standard techniques' [6,7] in the presence of glycerol. Thus, glycerol can be added to stabilize the tertiary and quaternary structure of large, multisubunit men> branc proteins without interf'cring with crystallization.

M o r e from the electron transport chain: the bacterial cytochrome c oxidase 'l'hc Paraco~zns d~'nitr~/k~/n.~ [9] and beef heart [10,11] cvtochromc c oxidascs revealed a conserved structurc for the catalytic core of st, bunits I and 11, which contain all the machincrv for thc reduction of oxvgcn and the pmnping of protons. T h c four sul)unit P. d~'nit/ffkzm.¢ enzyme offers many technical advantages for more dctailcd studies, although problems with crystal growth and diffraction anisntrnpy plagued the rcsearch [14]. Recently, ()stcrmeicr e'ta/. [15"1 rcportcd proccdures to grow well difTracting crystals of an actixc subunit l-subunit I1 core that avoids many of the problems encotmtered carlier. T h e importance of this work to the field of membrane protein structural biology is the demonstration of how refined methods for purit~'ing and crystallizing membrane protcins haxc become. T h c t w o sUbtll]it cvtochrome g oxidase core ~as prepared as a complex with a recombinant monoclonal F, antibody fragment previously engineered to help mediate crystallization [14l. T h e purificd F~-enzyme complex was then screened for crystallizability in the presence of different detergents. T h e resulting crystals revealed that, while the strategy of using a l:~.-cnz~me complex worked again, the molecular packing in the crystal was completely different. In the original crystals of the fi)ur-subunit enzyme, the molecular contacts between the F , - e n z y m e complexes were mcdiatcd entirely by the F~-fragment [141. In crystals of the two-subunit enzyme, the cytoplasmic and pcriplasmic surfaccs of the cnzvme n(m participate in crystal contacts. As might then be expected, thc crystallization of the two sul)unit cvtochrol-nC g oxidase core was more sensitive to the nature of thc dctergcnt. Ostcrmcier eta/. [1S'] found that two detergents allm~cd the growth of X-ra\ quality crystals: tmdccyl-~-l)-maltosidc and cych)heyxl-hex.vl-~-l)-maltoside. Interestingly; the latter detcrgent x~as designed by the

:345

company Anatrace (Maumec, ()hi(), lISA) specifically for membrane protein crystallization.

Bacteriorhodopsin revisited T h e reccnt structure determination of bactcriorhodopsin by X-ray crystallography [16"] is interesting in terms of a novel technical development in membrane protein crystallization. Despite its crystallization ahnost 19 years ago [2], the quality of the bacteriorhodopsin crystals did not allow the elucidation of its X-ray crystal structure. [!ntil recently, all of the high resolution structural work on bacteriorhodopsin was performed using electron crystallography on membranous two-dimensional crystals [17]. PebavPeyroula eta/. [16°], however, reported the X-ray crystal structure of bacteriorhodopsin using crystals grown from cubic mcsophases crcated by 'ternary' systems of water, detergcnt, solubilizcd protcin and lipid. Binary mixtt, res of lipids and water x~ill often display two or more mcsophases: the lipid bilavcr is the most common but cubic mesophascs also exist. T h e discontinuous or micellar cubic phase can bc easily visualized as the consequence of the cubic close-packing of lipid micclles. In contrast, the bicontint,ous cubic phase arises from the assembl.v of lipid monomers into a three-dimensional bilaver-like structure in which the lipid and aqueous phases are connected by channels. \Vhcrcas this lipidic structure does apply fixed spatial constraints as to where the membrane proteins can reside, l)ebay-Peyrouhl eta/. [16"] propose that there should be littic obstacle to the diffusion of mcmbrane proteins throughout the cubic mesot)hase created b v lipids. If a membranc protein could be added to such a lipid mesophase at high cnough concentration, the lipid mcsophase could act as a cr\stallization medium, nucleating cr.vstal growth within a more native, bilax/er-like cnvironmcnt. T h e resulting crystals of bacteriorhodopsin from a bicontinuous cubic lipid phase arc small but can display diffraction to 2.0 :~. l:urthermnre, the space group observed is P63 and, therefore, demonstrates that the bacteriorhodopsin moleculcs can not only mo~c within the lipid phase, but also adopt a packing arrangement that is not defined by the lipid phase. Hence, cubic lipid mcsophases can initiate membrane protein crystallization without limiting severely how the protein molecules will pack within the crystal. T h c refined X-ray structure of bacteriorhodnpsin provides new and additional details about the confl)rnlation of important side-chains and the existence of internal water molecules putatixely involved in proton pumping. From a technical standpoint, howexer, this work highlights the fact that some membrane proteins will crystallize without relying on a dctergcnt-solubilized sample [6,7].

Pores and more pores Structural biology of the bacterial outer membrane porins, a class of passive diffusion pores [18] that allow the passage of small molecules across the bilayer, is one of the more suctess fill areas of membrane protein rcsearch. T h e structure

346

Protein engineering

tff all known porins are rather simple: an antiparallel [3-barrcl creates the basic pore, while an 'eyelet' loop of polypeptide lining the inner barrel wall defines the characteristics of the pore. In the membrane, the monomcric pnrc usually associates into it symmetric trimer so that the pnrc axes align along the molecular threefold or perpendicular to the plane of the membrane. Porins are attractive model s y s t e m s to study passive transport for a number of rcasnns: firstly, it is easy to engineer mutant forms of them; secondly, they are physically robust: thirdly, transport is easy to measure in rcctmstitutcd systcms; and finally, they are relatively easy to crystallize. Attempts to understand the physical basis fl)r pore selectivity in the sugar-selective pnrins, such as maltoporins, haxe yielded more succcssfill rcsults [19"-21"]. T h e qucstion addressed hcre is how does a passive pore acting as a sizelimited molecular sieve affcct selectivc 'facilitated' diffusion of disaccharides and larger oligosaccharidcs through the pore. 'l'he structurc of l+;s~ezit'/zir/ (o/i maltnpnrin appeared in 1995 [22] and revealed a new porin chlss haxing a common 18-strandcd [~-barrel structure as opposed to the O m p F porin 16-stranded structure [18]. T h e structures of two othcr 5'. o'pkimuHum sugar-selective porins complcxed with a variety of ligands are now availablc [Z1",231. In their study, Wang eta/. [19 °j c[carh' show how thc disacchaddcs sucrose, trehalnse and melibiose interact with the pore. From this work, they can relate their structural observations to the individual sugar translocation ratcs in K. coil maltoporin: as the interaction of the saccharide units with the constriction loop i m p r m e s , the more cffective thc translocation appears to be. In contrast, Forst eta/. [21"] elucidated the sites of interaction for sucrose in the sucrosespecific pnrin ScrY and can propose that the difference in sucrnsc pcrmeation rates between Scr'l and maltoponn arise from differences in the topography and amino acid composition of thc respective pores. Meyer and Schulz [20"1 have a t t e m p t e d to go flirther with the 5'. o'pkimuHum maltodextrin-spccific porin and d e t e r m i n e thc probable e n e r g y profile along the permeation pathway With thesc efforts, it is possible for the first time to study thc physical nature of st)lute translocatit)n on an atomic level.

C h a n n e l s : a first l o o k ()ur knowledge of mammalian channel structure and function took at quantum leap forward with the structure determination of the 5'treptomvces /i~,Jdatzs KcsA K + channel I24"1, a prokaryotic analog of the cukaryotic K + channels. T h e KcsA K + channel is a m e m b e r of the two m e m b r a n e spanning segment, tetrameric K + channels. T h e KcsA K + channel protein, however, exhibits reginns of substantial s e q u e n c e homology to the six m e m b r a n e - s p a n n i n g segm e n t K + channels, suggesting that the smaller channels are truncated versions of their larger hnmologues. MacKinnon and co-wnrkers [24"1 were able to clone and overcxpress the KcsA K + channel, which is stablc in a detergent-sohibilized state, with a cleavable polyHis-tag as an aid for rapid purification. After purification, the protein was crystallized

in the detergent laurvl dimethx/laminc-N-oxidc b y v a p o r diffusion. "lb phasc the X-ray diffraction data, six dM'ercnt cystcine mtltants were prepared for heaxv atom dcrivatization; after multiplc isnllmrphous phasing, a combination ot d e n s i t \ lnodification and molecular a~eraging allowcd the calculation of electron density maps at 3.2 .~ rcsolution. A model of KcsA K + c h a n n c l , from residues 23-119, reveals a symmetric tetramcr with each monomer being composcd of two long rectal)rant-spanning c(-hcliccs and a short 'porc' helix (Figure 2). T h c pore is creatcd primarily by the surfaces of the 'inncr' and the pore helices. "l'hc Figure 2

Turret \

Selectivity filter

Pore heli~

-....

Outside

g c~

i

E

Outer helix

o

Inside

C u r r e n t O p i n i o n in

Biotechnology

A ribbon diagram of the KcsA potassium channel [24 "°] showing the general architecture of a single monomer (light gray) within the homotetramer.

selectivity filter is a narrow, 12 .~ long constriction created by one of the flanking p o l y p e p t i d c loops of the pore helix (Figurc 2); in fact, the positions of K + cations, ligated by m a i n - c h a i n carbnnyls, were c o n f i r m c d by d i f f c r c n c c 1;ourier analysis with crystals containing Rb + and Cs + ions. 'Fhe binding sites for the channel blockers tctracthvlammnnium [24"] and agitnxin 2 [25"] were also characterizcd. With this one structure, many of the physical principles hypothesizcd to account for selective K + conductance arc now confimlcd and a ncw era in structurc-bascd drug design for ion channels has begun. Eukaryntic channels are also beginning to reveal their structtlres and raising hopes for high resohition structures in the near future. Aquaporins (AQPs) are integral membrane proteins that t\lcilitate the transport of water across

Membrane protein structures Garavito 347

the cell membrane in eukaryotes and prokaryotes. The structure of AQP-I at around 6-7 A resolution has been determined by three different grot,ps [26"-28 °] t, sing electron microscopic analyses on two-dimensional AQP-1 crystals. AQP-1 is comprised of a right-handed barrel of six transmembrane helices. The center of thc helix bundle is believed to be the water pore and protein protuberances into this channel may define a selectivity region for water [26*]. These results reveal the power of modern electron microscopB a tcchniqne that is now limited by the technolngy for growing well-ordered two-dimensional crystals of membrane proteins [8"]. Interracial exist!

membrane

proteins:

they really do

The concept of an interracial membrane protein has undergone major changes over the past decade. This class of hybrid proteins does not include just peripheral membrane proteins, which bind to lipid bilavers via electrostatic interaction, but also membrane proteins that actually interact with, or even integrate into, the hydrophobic portion of the bilavcr [29,30]. [tnlike bitopic and polytopic membrane proteins, interfacial membrane protcins can either integrate transiently or permanently into membranes via hydrophobic interactions. In 1994, we reported on the structure of prostaglandin H svnthase (PGHS), the first integral membrane protein shown to be a monotopic, interracial membrane protein [29,31]: no

transmembrane segments appeared in the polypeptide chain. This raised the question as to how common were st,ch monotopic membrane proteins. The structure of squalene cvclase from A. m'idoca/arius [32 "°] st,ggests that it is another integral, but monotopic membrane protein and raises the possibility that such membrane proteins may be more common than originally believed. Squalene cyclasc, an integral membrane protein, converts squalene to hopene, a pentacyclic compound. The structure of detergent-solubilized squalene cyclase, at 2.9 A resolution, reveals a dimeric, bidomain protein. Domain 2 has a large internal, hydrophobic cavity which is presumed to be the activc site. Again, no transmembrane polypeptide segments are observed, but rather segments of polypeptide form a nonpolar 'plateau' (Figure 3). Even though no sequence or structural homology is observed between squalene cyclase and sheep PGHS, the squalene cyclase dimer orients these nonpolar plateaus in the monomers onto the same side of the direct, a feature that is highly reminiscent to the membrane-binding domains in the PGHS direct [29]. The polypeptide segments fnrming the nonpolar plateau also surround the mouth of the hydrophobic active-site cavity, a structural fcaturc also found in PGHS [31]. Hence, two enzymes that have evolved to act on apolar membrane components not only utilize the same strategy for membrane integration, but also the design of the active site entrances.

Figure 3

( "

(b)

Active site entrance

MBDs

1. . . . .

Active site entrance Current Opinion in Biotechnology

A comparison of the structures of (a) squalene cyclase and (b) ovine prostaglandin synthase show the relative locations of the membrane binding domains (MBDs), active site entrances, and bound ligand (dark gray) within the active sites.

348

Protein engineering

Many questions remain as to how interracial membrane proteins integrate into the bilayer. How dcep do they penetratc the bilaver? How much do they disturb local bilaver organization? Hermoso eta/. [33"] provide some insights into the kind on interactions that might occur at such a lipid-protein interface. Pancreatic lipases (Pl,s), which carry out the hydrolysis of triac>'lglyccrol, are laced with a problcnt of binding to emulsified lipid droplets in vivo. T h e efficient binding to the lipid droplets occurs bx, the formation of a complex between Pl, and a colipase (CI,). In the P I . - ( ; l , complex, the active site of PI. is exposed and a 50 :~ long hydrophobic surface is created. T h e crystal structure of the porcine P I . - ( ; l , complex [34], solved in the presence of detergents, suggested that the obscrved '()pen' and active c()nfonnation of the enzyme was stabilized by specific protein intcractions to the detergent phase within the crs'stal. Hermoso eta/. [33"'] dctermined the location and structure of the detergent phase using neutron diffraction and I ) 2 0 / H , O contrast variation, a technique used to study detergent-protein interactions in transmembrane proteins [35]. T h e concave face of CI. and the distal tip of the carboxy-terminal domain of PI, interact extensively with a discrete disk-shaped micellar region of detergcnt. Their resuhs suggest a novel mt, tli-step process of PI. activation: PI, and (;I, form a complex with a small lipid-st, rfactant micelle, the activc conformation of the enzyme is thus stabilized, and then the Pl.-Cl~-micelle complex integrates into the surface of a much larger lipid droplet.

Conclusions "l~echnological refinemcnts in preparing and crystallizing membrane proteins are now allowing a wide range of structures to be determined. This not only promises that new membrane protein structures will appear yearly, but also that more sophisticated and detailed structure-function analyses are now feasible.

Acknowledgements 1 would like to thank E Bcrr'y and R MacKinnon for providing mc with images of their protein structures that resulted in Figures 1 and 2, rcspcctivcl'y.

References and recommended reading Papers of particular interest, published within the annual period of review, have been highlighted as: • of special interest • * of outstanding interest 1.

Garavito R, Rosenbusch J: Three-dimensional crystals of an integral membrane protein: an initial X-ray analysis. J Ceil Biol 1980, 86:32?-329.

2.

Michel H, Oesterhelt D: Three-dimensional crystals of membrane proteins: bacteriorhodopsin. Proc Natl Acad Sci USA 1980, 77:1283-1285.

3.

Allen J P, Feher G, Yeates TO, Komiya K, Rees D: Structure of the reaction center from Rhodobacter sphaeroides R-26: the cofactors. Proc Nat Acad Sci USA 1987, 84:5730-5?34.

4.

Chang CH, Tied• D, Tang J, Smith U, Norris J, Schiffer M: Structure of Rhodopseudomonas sphaeroides R-26 reaction center. FEBS Lett 1986, 205:82-86.

5.

Henderson R, Unwin P: Three-dimensional model of purple membrane obtained by electron microscopy. Nature 1975, 257:28-32.

6.

Garavito RM, Picot D, Loll PJ: Strategies for crystallizing membrane proteins. J Bioenerg Biomembr 1996, 28:13-27.

?.

Ostermeier C, Michel H: Crystallization of membrane proteins. Curt Opin Struct Biol 1997, 7:697-701.

8. •

Heymann J, M011erD, force microscopy of 199'7, 7:543-549. A recent review covering atomic force microscopy structure. 9.

Mitsuoka K, Engel A: Electron and atomic membrane proteins. Curr Opin Struct Biol the current status of electron microscopy and as techniques to examine membrane protein

Iwata S, Ostermeier C, Ludwig B, Michel H: Structure at 2.8 A resolution of cytochrome c oxidase from Paracoccus denitrificans. Nature 1995, 376:660-669.

10. Tsukihara T, Aoyama H, Yamashita E, Tomizaki T, Yamaguchi H, Shinzawa-ltoh K, Nakashima R, Yaono R, Yoshikawa S: Structures of metal sites of oxidized bovine heart cytochrome c oxidase at 2.8 A, Science 1995, 269:1069-t 074. 11. Tsukihara T, Aoyama H, Yamashita E, Tomizaki T, Yamaguchi H, Shinzawa-ltoh K, Nakashima R, Yaono R, Yoshikawa S: The whole structure of the 13-subunit oxidized cytochrome c oxidase at 2.8 .~.. Science 1996,272:1136-1144. 12. Xia D, Yu CA, Kim H, Xia JZ, Kachurin AM, Zhang L, Yu L, •Deisenhofer J: Crystal structure of the cytochrome bcl complex from bovine heart mitochondria. Science 199?, 277:60-66. The work of Xia et aL and that of Zhang et aL [13"] represents not only the first structural characterizations of the cytochrome bc 1 complex, but also provide enough information on the tertiary structure to provide explanations of its function. 13. Zhang Z, Huang L, Shulmeister V, Chi Y-I, Kim K, Crofts A, Berry E, • * Kim S-H: Electron transfer by domain movement in cytochrome be1. Nature 1998, 392:677-684. The structural comparisions between eytochrome bc 1 complexes in different oxidation and ligand states allows Zhang et aL to propose that domain movement in the Rieske iron-sulfur protein is involved in electron shuttling. 14. Ostermeier C, Iwata S, Ludwig B, Michel H: Fv fragment-mediated crystallization of the membrane protein bacterial cytochrome c oxidase. Nat Struct Biol 1995, 2:842-846. 15. Ostermeier C, Harrenga A, Ermler U, Michel H: Structure at 2.7 A • resolution of the Paracoccus denitrificans two-subunit cytochrome c oxidase complexed with an antibody FV fragment. Proc Nat/Acad Sci USA 1997, 94:10547-10553. A important example of applying techniques from molecular biology and protein chemistry to a problem in membrane protein crystallization. 16. Pebay-Peyroula E, Rummel G, Rosenbusch JP, Landau EM: X-ray • structure of bacteriorhodopsin at 2.5 angstroms from microcrystals grown in lipidic cubic phases. Science 1997, 277:1676-1681. The use of lipidic cubic phases to grow three-dimensional crystals of bacteriorhodopsin finally allowed the determination of its structure by X-ray crystallography. The application of this technique to other membrane protein systems might prove very promising. 1 ?. Grigorieff N, Ceska TA, Downing KH, Baldwin JM, Henderson R: Electron-crystallographic refinement of the structure of bacteriorhodopsin. J Mol Biol 1996, 259:393-421 18. Schulz G: Porins: general to specific, native to engineered passive pores. Curt Opin Struct Biol 1996, 6:485-490. 19. Wang YF, Dutzler R, Rizkallah PJ, Rosenbusch JP, Schirmer T: • Channel specificity: structural basis for sugar discrimination and differential flux rates in maltoporin. J Mol Biol 199?, 272:56-63. X-ray crystallographic analyses of E. coil maltoporin in the presence of different disaccharides reveal differences in the modes of sugar binding that can be directly correlated to sugar discrimination and permeation by maltoporin. 20. •

Meyer JE, Schulz GE: Energy profile of maltooligosaccharide permeation through maltoporin as derived from the structure and from a statistical analysis of saccharide-protein interactions. Protein Sci 1997, 6:1084-1091. The authors characterize the pore surface of a maltoporie from S. typhimurium in order to derive the energy profile for sugar translocation. 21. Forst D, Welte W, Wacker T, Diederichs K: Structure of the sucrose. specific porin ScrY from Salmonella typhimurium and it complex with sucrose. Nat Struct Biol 1998, 5:37-46. The structure of the sucrose-specific porin ScrY from S. typhimurium shows how this porin adapts the pore lining to allow better sucrose translocation than maltoDorin.

M e m b r a n e protein structures Garavito

22. Schirmer T, Keller T, Wang Y, Rosenbusch JP: Structural basis for sugar translocation through maltoporin channels at 3.1 .~, resolution. Science 1995, 267:512-514. 23.

Meyer JE, Hofnung M, Schulz GE: Structure of maltoporin from Salmonella typhimurium ligated with a nitrophenyl-maltotrioside. J Mol Biol 1997, 266:761 -??5.

24. •-

Doyle D, Cabral J, Pfuetzner R, Kuo A, Gulbis J, Cohen S, Chait B, MacKinnon R: The structure of the potassium channel: molecular basis of the K+ conduction and selectivity. Science 1998, 280:697?. The first structure of an ion channel that is directly relevant to neurophysiology. The prokaryotic potassium channel structure determined by Doyle et aL explains many of the fundamental observations of nerve physiology over the past few decades. 25. MacKinnon R, Cohen S, Kuo A, Lee A, Chait B: Structural • conservation in prokaryotic and eukaryotic potassium channels. Science 1998, 280:106-109. The authors clearly show that the prokaryotic potassium channel structure can be used as a model system to explore toxin binding behavior observed in eukaryotic potassium channels. 26. Cheng A, van Hoek AN, Yeager M, Verkman AS, Mitra AK: Three* dimensional organization of a human water channel. Nature 1997, 387:62?-630. Along with [27"] and [28"], the work descibed here provides the first look into the structure of a ubiquitous channel protein in vertebrates and invertebrates. Furthermore, these reports demonstrate the rapid progress of electron microscopic research on membrane proteins.

349

See annotation to [26°]. 29.

Picot D, Garavito R: Prostaglandin H synthase: implications for membrane structure, FEBS Lett 1994, 346:21-25.

30.

Hurley J, Grobler J: Protein kinase C and phospholipase C: bilayer interactions and regulation. Curt Opin Struct Biol 199?, 7:557-565.

31.

Picot D, Loll PJ, Garavito RM: The X-ray crystal structure of the membrane protein prostaglandin H2 synthase-l. Nature 1994, 367:243-249.

32. Wendt K, Poralla K, Schulz G: Structure and function of a squalene •. cyclase. Science 1997, 277:1811-1815. The second structure of an integral membrane protein that displays an interfacial, monotopic interaction with lipid bilayers. Furthermore, the relationship of the membrane-binding domain of squalene cyclase to its active sites is highly reminiscent to that of prostaglandin synthase [29,30]. This suggests that many more membrane-bound enzymes that act on lipid or apolar substrates may have interfacial, monotopic interaction with biomembranes. 33. •*

Hermoso J, Pignol D, Penel S, Roth M, Chapus C, Fontecilla Camps JC: Neutron crystallographic evidence of lipase-colipase complex activation by a micelle. EMBO J 1997, 16:5531-5536. An important example of how single-crystal neutron crystallography can provide clear structural information about how interracial, monotopic interactions may occur between lipids and proteins. The pancreatic lipase-colipase complex structure reveal interracial, interactions with a detergent micelle which may then create the active hydrophobic binding surface that directly binds to lipid droplets.

27. •

Walz T, Hirai T, Murata K, Heymann JB, Mitsuoka K, Fujiyoshi Y, Smith BL, Agre P, Engel A: The three-dimensional structure of aquaporin-l. Nature 1997, 387:624-627. See annotation to [26"].

34.

Hermoso J, Pignol D, Kerfelec B, Crenon I, Chapus C, Fontecilla Camps J: Lipase activation by nonionic detergents: the crystal structure of the porcine lipase-colipase-tetraoxyethylene monooctylether complex. J Biol Chem 1996, 271:18007-18016.

28. Li H, Lee S, Jap BK: Molecular design of aquaporin-1 water • channel as revealed by electron crystallography. Nat Struct Biol 199?, 4:263-265.

35.

Pebay-Peyroula E, Garavito R, Rosenbusch J, Zulauf M, Timmins P: Detergent structure in tetragonal crystals of porin from the outer membrtane of E. coil Structure 1995, 3:1051-1059.