X-ray structure of a protein-conducting channel

A conserved heterotrimeric membrane protein complex, the Sec61 or SecY complex, forms a protein-conducting channel, allowing polypeptides to be transferred across or integrated into membranes. We report the crystal structure of the complex from Methanococcus jannaschii at a resolution of 3.2 A. The structure suggests that one copy of the heterotrimer serves as a functional translocation channel. The alpha-subunit has two linked halves, transmembrane segments 1-5 and 6-10, clamped together by the gamma-subunit. A cytoplasmic funnel leading into the channel is plugged by a short helix. Plug displacement can open the channel into an 'hourglass' with a ring of hydrophobic residues at its constriction. This ring may form a seal around the translocating polypeptide, hindering the permeation of other molecules. The structure also suggests mechanisms for signal-sequence recognition and for the lateral exit of transmembrane segments of nascent membrane proteins into lipid, and indicates binding sites for partners that provide the driving force for translocation.     

X-ray structure of a DNA hairpin molecule

We have solved the crystal structure of a synthetic DNA hexadecanucleotide of sequence: C-G-C-G-C-G-T-T-T-T-C-G-C-G-C-G, at 2.1 A resolution, and observed that it adopts a monomeric hairpin configuration with a Z-DNA hexamer stem. In the T4 loop the bases stack with one another and with neighbouring molecules of the crystal, and not with base pairs of their own hexamer stem. Two thymine T10 rings from different molecules stack between the C1-G16 ends of a third and a fourth hairpin helix, in a manner that suggests T-T base 'pairing' and simulates a long, 13-base-pair helix. Although such T-T interactions would not be present in solution, they illustrate a remarkable tendency of thymines for self-association. Purine-purine G-A base pairs are known to exist in the anti-anti conformation with an increase in local helix width; it may be that more serious consideration should be given to the possible existence of pyrimidine-pyrimidine C-T base pairs with decreased local helix width, particularly where several such base pairs occur sequentially.     

X-ray structure of a voltage-dependent K+ channel

Voltage-dependent K+ channels are members of the family of voltage-dependent cation (K+, Na+ and Ca2+) channels that open and allow ion conduction in response to changes in cell membrane voltage. This form of gating underlies the generation of nerve and muscle action potentials, among other processes. Here we present the structure of KvAP, a voltage-dependent K+ channel from Aeropyrum pernix. We have determined a crystal structure of the full-length channel at a resolution of 3.2 A, and of the isolated voltage-sensor domain at 1.9 A, both in complex with monoclonal Fab fragments. The channel contains a central ion-conduction pore surrounded by voltage sensors, which form what we call 'voltage-sensor paddles'-hydrophobic, cationic, helix-turn-helix structures on the channel's outer perimeter. Flexible hinges suggest that the voltage-sensor paddles move in response to membrane voltage changes, carrying their positive charge across the membrane.     

X-ray structure of a prokaryotic pentameric ligand-gated ion channel

Pentameric ligand-gated ion channels (pLGICs) are key players in the early events of electrical signal transduction at chemical synapses. The family codes for a structurally conserved scaffold of channel proteins that open in response to the binding of neurotransmitter molecules. All proteins share a pentameric organization of identical or related subunits that consist of an extracellular ligand-binding domain followed by a transmembrane channel domain. The nicotinic acetylcholine receptor (nAChR) is the most thoroughly studied member of the pLGIC family (for recent reviews see refs 1-3). Two sources of structural information provided an architectural framework for the family. The structure of the soluble acetylcholine-binding protein (AChBP) defined the organization of the extracellular domain and revealed the chemical basis of ligand interaction. Electron microscopy studies of the nAChR from Torpedo electric ray have yielded a picture of the full-length protein and have recently led to the interpretation of an electron density map at 4.0 A resolution. Despite the wealth of experimental information, high-resolution structures of any family member have so far not been available. Until recently, the pLGICs were believed to be only expressed in multicellular eukaryotic organisms. The abundance of prokaryotic genome sequences, however, allowed the identification of several homologous proteins in bacterial sources. Here we present the X-ray structure of a prokaryotic pLGIC from the bacterium Erwinia chrysanthemi (ELIC) at 3.3 A resolution. Our study reveals the first structure of a pLGIC at high resolution and provides an important model system for the investigation of the general mechanisms of ion permeation and gating within the family.     

Crystal structure and mechanism of a bacterial fluorinating enzyme

Fluorine is the thirteenth most abundant element in the earth's crust, but fluoride concentrations in surface water are low and fluorinated metabolites are extremely rare. The fluoride ion is a potent nucleophile in its desolvated state, but is tightly hydrated in water and effectively inert. Low availability and a lack of chemical reactivity have largely excluded fluoride from biochemistry: in particular, fluorine's high redox potential precludes the haloperoxidase-type mechanism used in the metabolic incorporation of chloride and bromide ions. But fluorinated chemicals are growing in industrial importance, with applications in pharmaceuticals, agrochemicals and materials products. Reactive fluorination reagents requiring specialist process technologies are needed in industry and, although biological catalysts for these processes are highly sought after, only one enzyme that can convert fluoride to organic fluorine has been described. Streptomyces cattleya can form carbon-fluorine bonds and must therefore have evolved an enzyme able to overcome the chemical challenges of using aqueous fluoride. Here we report the sequence and three-dimensional structure of the first native fluorination enzyme, 5'-fluoro-5'-deoxyadenosine synthase, from this organism. Both substrate and products have been observed bound to the enzyme, enabling us to propose a nucleophilic substitution mechanism for this biological fluorination reaction.     

X-ray structure of phospholipase A2 complexed with a substrate-derived inhibitor

Phospholipases A2 play a part in a number of physiologically important cellular processes such as inflammation, blood platelet aggregation and acute hypersensitivity. These processes are all initiated by the release of arachidonic acid from cell membranes which is catalysed by intracellular phospholipases A2 and followed by conversion of arachidonic acid to prostaglandins, leukotrienes or thromboxanes. An imbalance in the production of these compounds can lead to chronic inflammatory diseases such as rheumatoid arthritis and asthma. Inhibitors of phospholipase A2 might therefore act to reduce the effects of inflammation, so structural information about the binding of phospholipase A2 to its substrates could be helpful in the design of therapeutic drugs. The three-dimensional structure is not known for any intracellular phospholipase A2, but these enzymes share significant sequence homology with secreted phospholipases, for which some of the structures have been determined. Here we report the structure of a complex between an extracellular phospholipase A2 and a competitively inhibiting substrate analogue, which reveals considerable detail about the interaction and suggests a mechanism for catalysis by this enzyme.     

Crystal structure of a bacterial homologue of Na+/Cl--dependent neurotransmitter transporters

Na+/Cl--dependent transporters terminate synaptic transmission by using electrochemical gradients to drive the uptake of neurotransmitters, including the biogenic amines, from the synapse to the cytoplasm of neurons and glia. These transporters are the targets of therapeutic and illicit compounds, and their dysfunction has been implicated in multiple diseases of the nervous system. Here we present the crystal structure of a bacterial homologue of these transporters from Aquifex aeolicus, in complex with its substrate, leucine, and two sodium ions. The protein core consists of the first ten of twelve transmembrane segments, with segments 1-5 related to 6-10 by a pseudo-two-fold axis in the membrane plane. Leucine and the sodium ions are bound within the protein core, halfway across the membrane bilayer, in an occluded site devoid of water. The leucine and ion binding sites are defined by partially unwound transmembrane helices, with main-chain atoms and helix dipoles having key roles in substrate and ion binding. The structure reveals the architecture of this important class of transporter, illuminates the determinants of substrate binding and ion selectivity, and defines the external and internal gates.     

6 A-resolution X-ray structure of a variable surface glycoprotein from Trypanosoma brucei

The variable surface glycoprotein (VSG) is the predominant component of the surface coat of the African trypanosome. The expression of antigenically distinct VSGs on minor populations during infection allows the parasite to escape the host immune response. Purification of the protein is facilitated by the enzymatic release of a soluble form of VSG (sVSG) which occurs on cell lysis. The soluble form is a dimer with an approximate molecular weight of 120,000-130,000. Partial proteolysis of sVSG reveals a protease-sensitive link between an amino-terminal domain which comprises about two-thirds of the molecule, and a C-terminal domain which contains the membrane attachment site. We have obtained crystals suitable for high-resolution structural analysis from preparations of three sVSG: MITat 1.2, ILTat 1.25 and ILTat 1.22. The crystal structure of the dimer of the MITat 1.2 amino-terminal domain has been solved to 6 A resolution. We report here that the dimer is an unusual 90 A rod-like molecule composed of a helical bundle of at least four 80 A-long alpha-helices.     

Synthesis and X-ray crystal structure of a new manganese 18-metallacrown-6

0　IntroductionThemetalionsinmetallamacrocylesformedbysupramolecularself assemblycantaketrigonal,square planar,andtetrahe dralconfiguration[1 ,2 ] .Metallacrownisaspecialclassofmetalla macrocyle,theyareanalogoustocrownethersinbothstructureandfunctionexceptthatthemetalionsarenowtakingthepositionsofcoordinationatomsincrownether[3,4] .Themetallacrownswithdifferentnumberof [M—N—O]repeatunithavedifferentcavitysizes,suchas 9 MC 3[5 7] ,12 MC 4 [4,8 1 4 ] and 15 MC 5 [1 5] .Thereportedazameta…     

X-ray structure of a pentameric ligand-gated ion channel in an apparently open conformation

Pentameric ligand-gated ion channels from the Cys-loop family mediate fast chemo-electrical transduction, but the mechanisms of ion permeation and gating of these membrane proteins remain elusive. Here we present the X-ray structure at 2.9 A resolution of the bacterial Gloeobacter violaceus pentameric ligand-gated ion channel homologue (GLIC) at pH 4.6 in an apparently open conformation. This cationic channel is known to be permanently activated by protons. The structure is arranged as a funnel-shaped transmembrane pore widely open on the outer side and lined by hydrophobic residues. On the inner side, a 5 A constriction matches with rings of hydrophilic residues that are likely to contribute to the ionic selectivity. Structural comparison with ELIC, a bacterial homologue from Erwinia chrysanthemi solved in a presumed closed conformation, shows a wider pore where the narrow hydrophobic constriction found in ELIC is removed. Comparative analysis of GLIC and ELIC reveals, in concert, a rotation of each extracellular beta-sandwich domain as a rigid body, interface rearrangements, and a reorganization of the transmembrane domain, involving a tilt of the M2 and M3 alpha-helices away from the pore axis. These data are consistent with a model of pore opening based on both quaternary twist and tertiary deformation.     

Crystal structure of a bacterial homologue of the bile acid sodium symporter ASBT

High cholesterol levels greatly increase the risk of cardiovascular disease. About 50 per cent of cholesterol is eliminated from the body by its conversion into bile acids. However, bile acids released from the bile duct are constantly recycled, being reabsorbed in the intestine by the apical sodium-dependent bile acid transporter (ASBT, also known as SLC10A2). It has been shown in animal models that plasma cholesterol levels are considerably lowered by specific inhibitors of ASBT, and ASBT is thus a target for hypercholesterolaemia drugs. Here we report the crystal structure of a bacterial homologue of ASBT from Neisseria meningitidis (ASBT(NM)) at 2.2??. ASBT(NM) contains two inverted structural repeats of five transmembrane helices. A core domain of six helices harbours two sodium ions, and the remaining four helices pack in a row to form a flat, 'panel'-like domain. Overall, the architecture of the protein is remarkably similar to the sodium/proton antiporter NhaA, despite having no detectable sequence homology. The ASBT(NM) structure was captured with the substrate taurocholate present, bound between the core and panel domains in a large, inward-facing, hydrophobic cavity. Residues near this cavity have been shown to affect the binding of specific inhibitors of human ASBT. The position of the taurocholate molecule, together with the molecular architecture, suggests the rudiments of a possible transport mechanism.     

Liquid structure of pure iron by X-ray diffraction

The liquid structure of pure iron at 1540, 1560 and 1580℃ was studied by X-ray diffraction. The results show that near the melting point there is a medium-range order structure that fades away with the increasing temperature. The average nearest distance of atoms is almost independent of the melts temperature, but the average coordination number, the atom cluster size and the atom number in an atom cluster all decrease with the increasing temperature of the melt. Near the melting point there area lot of atom clusters in the pure iron melt. The atom cluster of pure iron has the body-centered cubic lattices, which are kept from the solid state. And the body-centered cubic lattices connect into network by occupying a same edge. The atoms in the surrounding of the atom clusters are arranged disorderly.     

Comparative crystal structure determination of griseofulvin: Powder X-ray diffraction versus single-crystal X-ray diffraction

In an attempt to compare crystal structure determination from powder data and single-crystal data,crystal structure of griseofulvin(C 17 H 17 ClO 6) was tested by both powder and single-crystal X-ray diffraction.Lattice parameters of griseofulvin are α=90.0°,a=b=8.9757,c=19.9345,V=1605.99 3 from powder data coinciding with α=90.0°,a=b=8.9714,c=19.8848,V=1600.46 3 from single-crystal data.Main processes of structure elucidating of griseofulvin by the two approaches were analyzed.Powder X-ray diffraction was demonstrated to be a powerful auxiliary implement to single-crystal X-ray diffraction in structure characterization,and its application can be popularized in the field of structure research of small organic molecules.     

Crystal structure of bacterial multidrug efflux transporter AcrB

AcrB is a major multidrug exporter in Escherichia coli. It cooperates with a membrane fusion protein, AcrA, and an outer membrane channel, TolC. We have determined the crystal structure of AcrB at 3.5 A resolution. Three AcrB protomers are organized as a homotrimer in the shape of a jellyfish. Each protomer is composed of a transmembrane region 50 A thick and a 70 A protruding headpiece. The top of the headpiece opens like a funnel, where TolC might directly dock into AcrB. A pore formed by three alpha-helices connects the funnel with a central cavity located at the bottom of the headpiece. The cavity has three vestibules at the side of the headpiece which lead into the periplasm. In the transmembrane region, each protomer has twelve transmembrane alpha-helices. The structure implies that substrates translocated from the cell interior through the transmembrane region and from the periplasm through the vestibules are collected in the central cavity and then actively transported through the pore into the TolC tunnel.     

Three-dimensional structure of the bacterial protein-translocation complex SecYEG

Transport and membrane integration of polypeptides is carried out by specific protein complexes in the membranes of all living cells. The Sec transport path provides an essential and ubiquitous route for protein translocation. In the bacterial cytoplasmic membrane, the channel is formed by oligomers of a heterotrimeric membrane protein complex consisting of subunits SecY, SecE and SecG. In the endoplasmic reticulum membrane, the channel is formed from the related Sec61 complex. Here we report the structure of the Escherichia coli SecYEG assembly at an in-plane resolution of 8 A. The three-dimensional map, calculated from two-dimensional SecYEG crystals, reveals a sandwich of two membranes interacting through the extensive cytoplasmic domains. Each membrane is composed of dimers of SecYEG. The monomeric complex contains 15 transmembrane helices. In the centre of the dimer we observe a 16 x 25 A cavity closed on the periplasmic side by two highly tilted transmembrane helices. This may represent the closed state of the protein-conducting channel.     

Design optimization of a C-band traveling-wave accelerating structure for a compact X-ray Free Electron Laser facility

The research and development of a C-band (5712 MHz) high gradient traveling-wave accelerating structure has been in progress for some years at the Shanghai Institute of Applied Physics, Chinese Academy of Sciences. Conceptual design of the accelerating structure has been accomplished, and verified by cold testing an experimental model. The first prototype structure is now ready for high RF power tests. To develop a robust high gradient C-band accelerating structure, an optimized constant gradient structure with a 4 /5 operating mode for future X-ray Free Electron Laser facilities is proposed in this paper. This scheme can improve the breakdown stability and beam quality. The high power RF system, four operating modes and disk iris shape are analyzed and optimized.