Structure of fumarate reductase from Wolinella succinogenes at 2.2 A resolution

Fumarate reductase couples the reduction of fumarate to succinate to the oxidation of quinol to quinone, in a reaction opposite to that catalysed by the related complex II of the respiratory chain (succinate dehydrogenase). Here we describe the crystal structure at 2.2 A resolution of the three protein subunits containing fumarate reductase from the anaerobic bacterium Wolinella succinogenes. Subunit A contains the site of fumarate reduction and a covalently bound flavin adenine dinucleotide prosthetic group. Subunit B contains three iron-sulphur centres. The menaquinol-oxidizing subunit C consists of five membrane-spanning, primarily helical segments and binds two haem b molecules. On the basis of the structure, we propose a pathway of electron transfer from the dihaem cytochrome b to the site of fumarate reduction and a mechanism of fumarate reduction. The relative orientations of the soluble and membrane-embedded subunits of succinate:quinone oxidoreductases appear to be unique.     

An unusual feature revealed by the crystal structure at 2.2 A resolution of human transforming growth factor-beta 2.

Transforming growth factor type beta 2 (TGF-beta 2) is a member of an expanding family of growth factors that regulate proliferation and differentiation of many different cell types. TGF-beta 2 binds to various receptors, one of which was shown to be a serine/threonine kinase. TGF-beta 2 is involved in wound healing, bone formation and modulation of immune functions. We report here the crystal structure of TGF-beta 2 at 2.2 A resolution, which reveals a novel monomer fold and dimer association. The monomer consists of two antiparallel pairs of beta-strands forming a flat curved surface and a separate, long alpha-helix. The disulphide-rich core has one disulphide bone pointing through a ring formed by the sequence motifs Cys-Ala-Gly-Ala-Cys and Cys-Lys-Cys, which are themselves connected through the cysteines. Two monomers are connected through a single disulphide bridge and associate such that the helix of one subunit interacts with the concave beta-sheet surface of the other. Four exposed loop regions might determine receptor specificity. The structure provides a suitable model for the TGF-beta s and other members of the super-family and is the basis for the analysis of the TGF-beta 2 interactions with the receptor.     

Structure of the C2 domain of human factor VIII at 1.5 A resolution

Human factor VIII is a plasma glycoprotein that has a critical role in blood coagulation. Factor VIII circulates as a complex with von Willebrand factor. After cleavage by thrombin, factor VIIIa associates with factor IXa at the surface of activated platelets or endothelial cells. This complex activates factor X (refs 6, 7), which in turn converts prothrombin to thrombin in the presence of factor Va (refs 8, 9). The carboxyl-terminal C2 domain of factor VIII contains sites that are essential for its binding to von Willebrand factor and to negatively charged phospholipid surfaces. Here we report the structure of human factor VIII C2 domain at 1.5 A resolution. The structure reveals a beta-sandwich core, from which two beta-turns and a loop display a group of solvent-exposed hydrophobic residues. Behind the hydrophobic surface lies a ring of positively charged residues. This motif suggests a mechanism for membrane binding involving both hydrophobic and electrostatic interactions. The structure explains, in part, mutations in the C2 region of factor VIII that lead to bleeding disorders in haemophilia A.     

Structure of the reovirus core at 3.6 A resolution

The reovirus core is an assembly with a relative molecular mass of 52 million that synthesizes, modifies and exports viral messenger RNA. Analysis of its structure by X-ray crystallography shows that there are alternative, specific and completely non-equivalent contacts made by several surfaces of two of its proteins; that the RNA capping and export apparatus is a hollow cylinder, which probably sequesters its substrate to ensure completion of the capping reactions; that the genomic double-stranded RNA is coiled into concentric layers within the particle; and that there is a protein shell that appears to be common to all groups of double-stranded RNA viruses.     

类风湿关节炎灭活滑膜液单个核细胞对非灭活滑膜液单个核细胞分泌IL-10水平的影响

目的观察类风湿关节炎灭活滑膜液单个核细胞对非灭活滑膜液单个核细胞分泌IL-10水平的影响。方法抽取类风湿关节炎(RA)患者关节液,分离关节液单个核细胞,分为对照组:2×10~6的非灭活单个核细胞,培养条件为37℃,5%CO_2孵育箱培养48h;实验组:细胞数为2×10~6个非灭活单个核细胞与甲醛灭活后的单个核细胞按1:10混合培养.用ELISA法测定培养上清中IL-10的水平.结果实验组较对照组IL-10水平明显升高(P<0.01).结论灭活滑膜液单个核细胞对非灭活滑膜液单个核细胞IL-10分泌有促进作用,提示未经克隆选择的RA滑膜液混合细胞可能具有诱导分泌IL-10CD_4调节性T细胞的生成作用.     

Structure of the nucleosome core particle at 7 A resolution

The crystal structure of the nucleosome core particle has been solved to 7 A resolution. The right-handed B-DNA superhelix on the outside contains several sharp bends and makes numerous interactions with the histone octamer within. The central turn of superhelix and H3 . H4 tetramer have dyad symmetry, but the H2A . H2B dimers show departures due to interparticle associations.     

Structure of the calcium regulatory muscle protein troponin-C at 2.8 A resolution

Crystals of turkey skeletal muscle troponin-C reveal a molecule of two domains with an unusual structure. Two Ca2+ ions are bound to the C-terminal domain. The two cation-binding sites of the regulatory (N-terminal) domain are Ca2+ free; this domain adopts a markedly different conformation from the C-terminal domain. The two domains are connected by a long nine-turn alpha-helix; three of these turns are exposed fully to solvent.     

Structure of a bacterial 30S ribosomal subunit at 5.5 A resolution.

The 30S ribosomal subunit binds messenger RNA and the anticodon stem-loop of transfer RNA during protein synthesis. A crystallographic analysis of the structure of the subunit from the bacterium Thermus thermophilus is presented. At a resolution of 5.5 A, the phosphate backbone of the ribosomal RNA is visible, as are the alpha-helices of the ribosomal proteins, enabling double-helical regions of RNA to be identified throughout the subunit, all seven of the small-subunit proteins of known crystal structure to be positioned in the electron density map, and the fold of the entire central domain of the small-subunit ribosomal RNA to be determined.     

Structure of acid-sensing ion channel 1 at 1.9 A resolution and low pH

Acid-sensing ion channels (ASICs) are voltage-independent, proton-activated receptors that belong to the epithelial sodium channel/degenerin family of ion channels and are implicated in perception of pain, ischaemic stroke, mechanosensation, learning and memory. Here we report the low-pH crystal structure of a chicken ASIC1 deletion mutant at 1.9 A resolution. Each subunit of the chalice-shaped homotrimer is composed of short amino and carboxy termini, two transmembrane helices, a bound chloride ion and a disulphide-rich, multidomain extracellular region enriched in acidic residues and carboxyl-carboxylate pairs within 3 A, suggesting that at least one carboxyl group bears a proton. Electrophysiological studies on aspartate-to-asparagine mutants confirm that these carboxyl-carboxylate pairs participate in proton sensing. Between the acidic residues and the transmembrane pore lies a disulphide-rich 'thumb' domain poised to couple the binding of protons to the opening of the ion channel, thus demonstrating that proton activation involves long-range conformational changes.     

Isolation of CD4- CD8- mycobacteria-reactive T lymphocyte clones from rheumatoid arthritis synovial fluid

The majority of peripheral T cells express a heterodimeric, alpha/beta T-cell receptor, which recognizes specific antigenic peptides bound to self major histocompatibility complex (MHC) molecules, and either the CD4 or CD8 surface markers. An additional subset of T cells, whose physiological function is unknown, express a distinct CD3-associated receptor composed of gamma and delta chains. This subset includes cells lacking both CD4 and CD8 surface markers, which may be involved in autoimmunity. The recognition specificity of the gamma/delta receptors is not well characterized and has been defined in only one case to date, a murine cell line which shows MHC-linked specificity. In this report, we describe the isolation of CD4- CD8-, gamma/delta TCR bearing T cell clones from the synovial fluid of a rheumatoid arthritis patient. These T cell clones respond specifically to mycobacterial antigens without MHC restriction.     

Structure of DNase I at 2.0 A resolution suggests a mechanism for binding to and cutting DNA

Bovine pancreatic deoxyribonuclease I (DNase I), an endonuclease that degrades double-stranded DNA in a nonspecific but sequence-dependent manner, has been used as a biochemical tool in various reactions, in particular as a probe for the structure of chromatin and for the helical periodicity of DNA on the nucleosome and in solution. Limited digestion by DNase I, termed DNase I 'footprinting', is routinely used to detect protected regions in DNA-protein complexes. Recently, we have solved the three-dimensional structure of this glycoprotein (relative molecular mass 30,400) by X-ray structure analysis at 2.5 A resolution and have subsequently refined it crystallographically at 2.0 A. Based on the refined structure and the binding of Ca2+-thymidine 3',5'-diphosphate (Ca-pTp) at the active site, we propose a mechanism of action and present a model for the interaction of DNase I with double-stranded DNA that involves the binding of an exposed loop region in the minor groove of B-DNA and electrostatic interactions of phosphates from both strands with arginine and lysine residues on either side of this loop. We explain DNase I cleavage patterns in terms of this model and discuss the consequences of the extended DNase I-DNA contact region for the interpretation of DNase I footprinting results.