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排序方式: 共有203条查询结果,搜索用时 31 毫秒
31.
Anneke Kuipers Jenny Wierenga Rick Rink Leon D. Kluskens Arnold J. M. Driessen Oscar P. Kuipers Gert N. Moll 《Applied microbiology》2006,72(12):7626-7633
Nisin is a lanthionine-containing antimicrobial peptide produced by Lactococcus lactis. Its (methyl)lanthionines are introduced by two posttranslational enzymatic steps involving the dehydratase NisB, which dehydrates serine and threonine residues, and the cyclase NisC, which couples these dehydrated residues to cysteines, yielding thioether-bridged amino acids called lanthionines. The prenisin is subsequently exported by the ABC transporter NisT and extracellularly processed by the peptidase NisP. L. lactis expressing the nisBTC genes can modify and secrete a wide range of nonlantibiotic peptides. Here we demonstrate that in the absence of NisT and NisC, the Sec pathway of L. lactis can be exploited for the secretion of dehydrated variants of therapeutic peptides. Furthermore, posttranslational modifications by NisB and NisC still occur even when the nisin leader is preceded by a Sec signal peptide or a Tat signal peptide 27 or 44 amino acids long, respectively. However, transport of fully modified prenisin via the Sec pathway is impaired. The extent of NisB-mediated dehydration could be improved by raising the intracellular concentration NisB or by modulating the export efficiency through altering the signal sequence. These data demonstrate that besides the traditional lantibiotic transporter NisT, the Sec pathway with an established broad substrate range can be utilized for the improved export of lantibiotic enzyme-modified (poly)peptides. 相似文献
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33.
Structural determinants for ligand binding and catalysis of triosephosphate isomerase. 总被引:1,自引:0,他引:1
I Kursula S Partanen A M Lambeir D M Antonov K Augustyns R K Wierenga 《European journal of biochemistry》2001,268(19):5189-5196
The crystal structure of leishmania triosephosphate isomerase (TIM) complexed with 2-(N-formyl-N-hydroxy)-aminoethyl phosphonate (IPP) highlights the importance of Asn11 for binding and catalysis. IPP is an analogue of the substrate D-glyceraldehyde-3-phosphate, and it is observed to bind with its aldehyde oxygen in an oxyanion hole formed by ND2 of Asn11 and NE2 of His95. Comparison of the mode of binding of IPP and the transition state analogue phosphoglycolohydroxamate (PGH) suggests that the Glu167 side chain, as well as the triose part of the substrate, adopt different conformations as the catalysed reaction proceeds. Comparison of the TIM-IPP and the TIM-PGH structures with other liganded and unliganded structures also highlights the conformational flexibility of the ligand and the active site, as well as the conserved mode of ligand binding. 相似文献
34.
Serena Donnini Alessandra Villa Gerrit Groenhof Alan E. Mark Rik K. Wierenga André H. Juffer 《Proteins》2009,76(1):138-150
When estimating binding affinities of a ligand, which can exists in multiple forms, for a target molecule, one must consider all possible competing equilibria. Here, a method is presented that estimates the contribution of the protonation equilibria of a ligand in solution to the measured or calculated binding affinity. The method yields a correction to binding constants that are based on the total concentration of inhibitor (the sum of all ionized forms of the inhibitor in solution) to account for the complexed form of the inhibitor only. The method is applied to the calculation of the difference in binding affinity of two inhibitors, 2‐phosphoglycolate (PGA) and its phoshonate analog 3‐phosphonopropionate (3PP), for the glycolytic enzyme triosephosphate isomerase. Both inhibitors have three titrating sites and exist in solution as a mixture of different forms. In this case the form that actually binds to the enzyme is present at relative low concentrations. The contributions of the alternative forms to the difference in binding energies is estimated by means of molecular dynamics simulations and corrections. The inhibitors undergo a pKa shift upon binding that is estimated by ab initio calculations. An interesting finding is that the affinity difference of the two inhibitors is not due to different interactions in the active site of the enzyme, but rather due to the difference in the solvation properties of the inhibitors. Protein 2009. © 2008 Wiley‐Liss, Inc. 相似文献
35.
The formation of a carbon-carbon bond is an essential step in the biosynthetic pathways by which fatty acids and polyketides are made. The thiolase superfamily enzymes catalyse this carbon-carbon-bond formation via a thioester-dependent Claisen-condensation-reaction mechanism. In this way, fatty-acid chains and polyketides are made by sequentially adding simple building blocks, such as acetate units, to the growing molecule. A common feature of these enzymes is a reactive cysteine residue that is transiently acylated in the catalytic cycle. The wide catalytic diversity of the thiolase superfamily enzymes is of great interest. In particular, the type-III polyketide synthases make complicated compounds of great biological importance using multiple, subsequent condensation reactions, which are all catalysed in the same active-site cavity. The crucial metabolic importance of the bacterial fatty-acid-synthesizing enzymes stimulates in-depth studies that aim to develop efficient anti-bacterial drugs. 相似文献
36.
BACKGROUND: Thiolases are ubiquitous and form a large family of dimeric or tetrameric enzymes with a conserved, five-layered alphabetaalphabetaalpha catalytic domain. Thiolases can function either degradatively, in the beta-oxidation pathway of fatty acids, or biosynthetically. Biosynthetic thiolases catalyze the biological Claisen condensation of two molecules of acetyl-CoA to form acetoacetyl-CoA. This is one of the fundamental categories of carbon skeletal assembly patterns in biological systems and is the first step in a wide range of biosynthetic pathways, including those that generate cholesterol, steroid hormones, and various energy-storage molecules. RESULTS: The crystal structure of the tetrameric biosynthetic thiolase from Zoogloea ramigera has been determined at 2.0 A resolution. The structure contains a striking and novel 'cage-like' tetramerization motif, which allows for some hinge motion of the two tight dimers with respect to each other. The protein crystals were flash-frozen after a short soak with the enzyme's substrate, acetoacetyl-CoA. A reaction intermediate was thus trapped: the enzyme tetramer is acetylated at Cys89 and has a CoA molecule bound in each of its active-site pockets. CONCLUSIONS: The shape of the substrate-binding pocket reveals the basis for the short-chain substrate specificity of the enzyme. The active-site architecture, and in particular the position of the covalently attached acetyl group, allow a more detailed reaction mechanism to be proposed in which Cys378 is involved in both steps of the reaction. The structure also suggests an important role for the thioester oxygen atom of the acetylated enzyme in catalysis. 相似文献
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Activation of a heterogeneous hepatitis B (HB) core and e antigen-specific CD4+ T-cell population during seroconversion to anti-HBe and anti-HBs in hepatitis B virus infection. 总被引:12,自引:5,他引:7 下载免费PDF全文
M C Jung H M Diepolder U Spengler E A Wierenga R Zachoval R M Hoffmann D Eichenlaub G Frsner H Will G R Pape 《Journal of virology》1995,69(6):3358-3368
Overcoming hepatitis B virus infection essentially depends on the appropriate immune response of the infected host. Among the hepatitis B virus antigens, the core (HBcAg) and e (HBeAg) proteins appear highly immunogenic and induce important lymphocyte effector functions. In order to investigate the importance of HBcAg/HBeAg-specific T lymphocytes in patients with acute and chronic hepatitis B and to identify immunodominant epitopes within the HBcAg/HBeAg, CD4+ T-cell responses to hepatitis B virus-encoded HBcAg and HBcAg/HBeAg-derived peptides were studied in 49 patients with acute and 39 patients with chronic hepatitis B. The results show a frequent antigen-specific CD4+ T-cell activation during acute hepatitis B infection, a rare HBcAg/HBeAg-specific CD4+ T-cell response among HBeAg+ chronic carriers, and no response in patients with anti-HBe+ chronic hepatitis. An increasing CD4+ T-cell response to HBcAg/HBeAg coincides with loss of HBeAg and hepatitis B virus surface antigen (HBsAg). Functional analysis of peptide-specific CD4+ T-cell clones revealed a heterogeneous population with respect to lymphokine production. Epitope mapping within the HBcAg/HBeAg peptide defined amino acids (aa) 1 to 25 and aa 61 to 85, irrespective of the HLA haplotype, as the predominant CD4+ T-cell recognition sites. Other important sequences could be identified in the amino-terminal part of the protein, aa 21 to 45, aa 41 to 65, and aa 81 to 105. The immunodominant epitopes are expressed in both proteins, HBcAg and HBeAg. Our findings lead to the conclusion that activation of CD4+ T lymphocytes by HBcAg/HBeAg is a prerequisite for viral elimination, and further studies have to focus on the question of how to enhance or induce this type of T-cell response in chronic carriers. The immunodominant viral sequences identified may have relevance to synthetic vaccine design and to the use of peptide T-cell sites as immunotherapeutic agents in chronic infection. 相似文献
39.
Coenzyme A is involved in a number of important metabolic pathways. Recently the structures of several coenzyme A binding proteins have been determined. We compare in some detail the structures of seven different coenzyme A protein complexes. These seven proteins all have distinctly different folds. 相似文献