Home About us Contact
|
Home About us Contact | ||
|
|
||
Substrate-binding Cleft (substrate-binding + cleft)
Selected AbstractsStructure and reactivity of Trypanosoma brucei pteridine reductase: inhibition by the archetypal antifolate methotrexateMOLECULAR MICROBIOLOGY, Issue 6 2006Alice Dawson Summary The protozoan Trypanosoma brucei has a functional pteridine reductase (TbPTR1), an NADPH-dependent short-chain reductase that participates in the salvage of pterins, which are essential for parasite growth. PTR1 displays broad-spectrum activity with pterins and folates, provides a metabolic bypass for inhibition of the trypanosomatid dihydrofolate reductase and therefore compromises the use of antifolates for treatment of trypanosomiasis. Catalytic properties of recombinant TbPTR1 and inhibition by the archetypal antifolate methotrexate have been characterized and the crystal structure of the ternary complex with cofactor NADP+ and the inhibitor determined at 2.2 Å resolution. This enzyme shares 50% amino acid sequence identity with Leishmania major PTR1 (LmPTR1) and comparisons show that the architecture of the cofactor binding site, and the catalytic centre are highly conserved, as are most interactions with the inhibitor. However, specific amino acid differences, in particular the placement of Trp221 at the side of the active site, and adjustment of the ,6-,6 loop and ,6 helix at one side of the substrate-binding cleft significantly reduce the size of the substrate binding site of TbPTR1 and alter the chemical properties compared with LmPTR1. A reactive Cys168, within the active site cleft, in conjunction with the C-terminus carboxyl group and His267 of a partner subunit forms a triad similar to the catalytic component of cysteine proteases. TbPTR1 therefore offers novel structural features to exploit in the search for inhibitors of therapeutic value against African trypanosomiasis. [source] Structure of BthA-I complexed with p -bromophenacyl bromide: possible correlations with lack of pharmacological activityACTA CRYSTALLOGRAPHICA SECTION D, Issue 12 2005Angelo J. Magro The crystal structure of an acidic phospholipase A2 isolated from Bothrops jararacussu venom (BthA-I) chemically modified with p -bromophenacyl bromide (BPB) has been determined at 1.85,Å resolution. The catalytic, platelet-aggregation inhibition, anticoagulant and hypotensive activities of BthA-I are abolished by ligand binding. Electron-density maps permitted unambiguous identification of inhibitor covalently bound to His48 in the substrate-binding cleft. The BthA-I,BPB complex contains three structural regions that are modified after inhibitor binding: the Ca2+ -binding loop, ,-wing and C-terminal regions. Comparison of BthA-I,BPB with two other BPB-inhibited PLA2 structures suggests that in the absence of Na+ ions at the Ca2+ -binding loop, this loop and other regions of the PLA2s undergo structural changes. The BthA-I,BPB structure reveals a novel oligomeric conformation. This conformation is more energetically and conformationally stable than the native structure and the abolition of pharmacological activities by the ligand may be related to the oligomeric structural changes. A residue of the `pancreatic' loop (Lys69), which is usually attributed as providing the anticoagulant effect, is in the dimeric interface of BthA-I,BPB, leading to a new hypothesis regarding the abolition of this activity by BPB. [source] Extraction of functional motion in trypsin crystal structuresACTA CRYSTALLOGRAPHICA SECTION D, Issue 8 2005Andrea Schmidt The analysis of anisotropic atomic displacement parameters for the direct extraction of functionally relevant motion from X-ray crystal structures of Fusarium oxysporum trypsin is presented. Several atomic resolution structures complexed with inhibitors or substrates and determined at different pH values and temperatures were investigated. The analysis revealed a breathing-like molecular motion conserved across trypsin structures from two organisms and three different crystal forms. Directional motion was observed suggesting a change of the width of the substrate-binding cleft and a change in the length of the specificity pocket. The differences in direction of motion across the structures are dependent on the mode of substrate or inhibitor binding and the chemical environment around the active-site residues. Together with the occurrence of multiple-residue conformers, they reflect spatial rearrangement throughout the deacylation pathway. [source] Structure of XynB, a highly thermostable ,-1,4-xylanase from Dictyoglomus thermophilum Rt46B.1, at 1.8,Å resolutionACTA CRYSTALLOGRAPHICA SECTION D, Issue 11 2000Andrew A. McCarthy Microorganisms employ a large array of enzymes to break down the cellulose and hemicelluloses of plant biomass. These enzymes, especially those with high thermal stability, have many uses in biotechnology. We have solved the crystal structure of a ,-1,4-xylanase, XynB, from the extremely thermophilic bacterium Dictyoglomus thermophilum, isolate Rt46B.1. The protein crystallized from 1.6,M ammonium sulfate, 0.2,M HEPES pH 7.2 and 10% glycerol, with unit-cell parameters a = b = 91.3, c = 44.9,Å and space group P43. The structure was solved at high resolution (1.8,Å) by X-ray crystallography, using the method of isomorphous replacement with a single mercury derivative, and refined to a final R factor of 18.3% (Rfree = 22.1%). XynB has the single-domain fold typical of family 11 xylanases, comprising a jelly roll of two highly twisted ,-sheets that create a deep substrate-binding cleft. The two catalytic residues, Glu90 and Glu180, occupy this cleft. Compared with other family 11 xylanases, XynB has a greater proportion of polar surface and has a slightly extended C-terminus that, combined with the extension of ,-strand A5, gives additional hydrogen bonding and hydrophobic packing. These factors may account for the enhanced thermal stability of the enzyme. [source] Structure of the complex of porcine pancreatic elastase with a trimacrocyclic peptide inhibitor FR901451ACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 9 2005Takayoshi Kinoshita Porcine pancreatic elastase (PPE) resembles the attractive drug target leukocyte elastase, which has the ability to degrade connective tissue in the body. The crystal structure of PPE complexed with a novel trimacrocyclic peptide inhibitor, FR901451, was solved at 1.9,Å resolution. The inhibitor occupied the subsites S3 through S3, of PPE and induced conformational changes in the side chains of Arg64 and Arg226, which are located at the edges of the substrate-binding cleft. Structural comparison of five PPE,inhibitor complexes, including the FR901451 complex and non-ligated PPE, reveals that the residues forming the S2, S1, S1, and S2, subsites in the cleft are rigid, but the two arginine residues playing a part in the S3 and S3, subsites are flexible. Structural comparison of PPE with human leukocyte elastase (HLE) implies that the inhibitor binds to HLE in a similar manner to the FR901451,PPE complex. This structural insight may help in the design of potent elastase inhibitors. [source] | ||||||||||