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Xiaoman Liu Louis A. Silks Prof. Dr. Cuiping Liu Dr. Morgane Ollivault‐Shiflett Xin Huang Jing Li Dr. Guimin Luo Prof. Dr. Ya‐Ming Hou Prof. Dr. Junqiu Liu Prof. Dr. Jiacong Shen Prof. 《Angewandte Chemie (International ed. in English)》2009,48(11):2020-2023
A rival to native peroxidase! An existing binding site for glutathione was combined with the catalytic residue tellurocysteine by using an auxotrophic expression system to create an engineered enzyme that functions as a glutathione peroxidase from the scaffold of a glutathione transferase (see picture). The catalytic activity of the telluroenzyme in the reduction of hydroperoxides by glutathione is comparable to that of native glutathione peroxidase.
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本文评述了天然蛋白质作为模拟酶的研究及生物活性物质分析中的应用进展。引用参考文献76篇。 相似文献
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An Off‐Pathway Folding Intermediate of an Acyl Carrier Protein Domain Coexists with the Folded and Unfolded States under Native Conditions 下载免费PDF全文
Dr. Jackwee Lim Tianshu Xiao Dr. Jingsong Fan Prof. Daiwen Yang 《Angewandte Chemie (International ed. in English)》2014,53(9):2358-2361
A protein can exist in multiple states under native conditions and those states with low populations are often critical to biological function and self‐assembly. To investigate the role of the minor states of an acyl carrier protein, NMR techniques were applied to determine the number of minor states and characterize their structures and kinetics. The acyl carrier protein from Micromonospora echinospora was found to exist in one major folded state (95.2 %), one unfolded state (4.1 %), and one intermediate state (0.7 %) under native conditions. The three states are in dynamic equilibrium and the intermediate state very likely adopts a native‐like structure and is an off‐pathway folding product. The intermediate state may mediate the formation of oligomers in vitro and play an important role in the recognition of partner enzymes in vivo. 相似文献
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Orit Weil-Ktorza Nischay Rege Shifra Lansky Deborah E. Shalev Prof. Gil Shoham Prof. Michael A. Weiss Prof. Norman Metanis 《Chemistry (Weinheim an der Bergstrasse, Germany)》2019,25(36):8513-8521
Insulin analogues, mainstays in the modern treatment of diabetes mellitus, exemplify the utility of protein engineering in molecular pharmacology. Whereas chemical syntheses of the individual A and B chains were accomplished in the early 1960s, their combination to form native insulin remains inefficient because of competing disulfide pairing and aggregation. To overcome these limitations, we envisioned an alternative approach: pairwise substitution of cysteine residues with selenocysteine (Sec, U). To this end, CysA6 and CysA11 (which form the internal intrachain A6–A11 disulfide bridge) were each replaced with Sec. The A chain[C6U, C11U] variant was prepared by solid-phase peptide synthesis; while sulfitolysis of biosynthetic human insulin provided wild-type B chain-di-S-sulfonate. The presence of selenium atoms at these sites markedly enhanced the rate and fidelity of chain combination, thus solving a long-standing challenge in chemical insulin synthesis. The affinity of the Se-insulin analogue for the lectin-purified insulin receptor was indistinguishable from that of WT-insulin. Remarkably, the thermodynamic stability of the analogue at 25 °C, as inferred from guanidine denaturation studies, was augmented (ΔΔGu ≈0.8 kcal mol−1). In accordance with such enhanced stability, reductive unfolding of the Se-insulin analogue and resistance to enzymatic cleavage by Glu-C protease occurred four times more slowly than that of WT-insulin. 2D-NMR and X-ray crystallographic studies demonstrated a native-like three-dimensional structure in which the diselenide bridge was accommodated in the hydrophobic core without steric clash. 相似文献
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Jaanus Kruusma Aidan Rhodes Radhika Bhatia J. A. Gareth Williams Adam M. Benham Ritu Kataky 《Journal of solution chemistry》2007,36(4):517-529
Increasing enrichment of dimethyl sulfoxide, DMSO, in DMSO-water mixtures causes a reversal in the thermodynamic dissociation
constants, pK
as, and has a marked effect on the redox potentails of the thiolic and amino groups in cysteine and the protein disulfide isomerase
(PDI) mimic BMC, Vectrase. This paper illustrates the effect of a hydrogen-bonding environment on the ionisation and redox
properties of thiol groups in amino acids. A combination of potentiometry and Raman spectroscopy was applied to rationalise
the observations. Intracellular environments are full of hydrophobic, hydrogen-bonding environments. The results illustrate
the profound effects of the local environment on the thiol group. 相似文献
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“Invisible” Conformers of an Antifungal Disulfide Protein Revealed by Constrained Cold and Heat Unfolding,CEST‐NMR Experiments,and Molecular Dynamics Calculations 下载免费PDF全文
Ádám Fizil Dr. Zoltán Gáspári Prof. Terézia Barna Prof. Florentine Marx Prof. Gyula Batta 《Chemistry (Weinheim an der Bergstrasse, Germany)》2015,21(13):5136-5144
Transition between conformational states in proteins is being recognized as a possible key factor of function. In support of this, hidden dynamic NMR structures were detected in several cases up to populations of a few percent. Here, we show by two‐ and three‐state analysis of thermal unfolding, that the population of hidden states may weight 20–40 % at 298 K in a disulfide‐rich protein. In addition, sensitive 15N‐CEST NMR experiments identified a low populated (0.15 %) state that was in slow exchange with the folded PAF protein. Remarkably, other techniques failed to identify the rest of the NMR “dark matter”. Comparison of the temperature dependence of chemical shifts from experiments and molecular dynamics calculations suggests that hidden conformers of PAF differ in the loop and terminal regions and are most similar in the evolutionary conserved core. Our observations point to the existence of a complex conformational landscape with multiple conformational states in dynamic equilibrium, with diverse exchange rates presumably responsible for the completely hidden nature of a considerable fraction. 相似文献
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Dr. Györgyi Váradi Prof. Gábor K. Tóth Dr. Zoltán Kele Dr. László Galgóczy Ádám Fizil Prof. Gyula Batta 《Chemistry (Weinheim an der Bergstrasse, Germany)》2013,19(38):12684-12692
The folding of disulfide proteins is of considerable interest because knowledge of this may influence our present understanding of protein folding. However, sometimes even the disulfide pattern cannot be unequivocally determined by the available experimental techniques. For example, the structures of a few small antifungal proteins (PAF, AFP) have been disclosed recently using NMR spectroscopy but with some ambiguity in the actual disulfide pattern. For this reason, we carried out the chemical synthesis of PAF. Probing different approaches, the oxidative folding of the synthetic linear PAF yielded a folded protein that has identical structure and antifungal activity as the native PAF. In contrast, unfolded linear PAF was inactive, a result that may have implications concerning its redox state in the mode of action. 相似文献
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Ccile Dubois Isaline Herrada Philippe Barthe Christian Roumestand 《Molecules (Basel, Switzerland)》2020,25(23)
High-hydrostatic pressure is an alternative perturbation method that can be used to destabilize globular proteins. Generally perfectly reversible, pressure exerts local effects on regions or domains of a protein containing internal voids, contrary to heat or chemical denaturant that destabilize protein structures uniformly. When combined with NMR spectroscopy, high pressure (HP) allows one to monitor at a residue-level resolution the structural transitions occurring upon unfolding and to determine the kinetic properties of the process. The use of HP-NMR has long been hampered by technical difficulties. Owing to the recent development of commercially available high-pressure sample cells, HP-NMR experiments can now be routinely performed. This review summarizes recent advances of HP-NMR techniques for the characterization at a quasi-atomic resolution of the protein folding energy landscape. 相似文献
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Lally Mitra Jean‐Baptiste Rouget Bertrand Garcia‐Moreno Prof. Dr. Catherine A. Royer Prof. Dr. Roland Winter Prof. Dr. 《Chemphyschem》2008,9(18):2715-2721
Herein, we probe by pressure perturbation calorimetry (PPC) the coefficient of thermal expansion, the volumetric and the hydration properties of variants of a hyperstable variant of staphylococcal nuclease (SNase), Δ+PHS. The temperature‐dependent volumetric properties of the folded and unfolded states of the wild‐type protein are calculated with previously published data. The present PPC results are used to interpret the volume diagram and expansivity at a molecular level. We conclude that the expansivity of the unfolded state is, to a first approximation, temperature independent, while that of the folded state decreases with increasing temperature. Our data suggest that at low temperature the defining contribution to ΔV comes mainly from excluded volume differences and ΔV for unfolding is negative. In contrast, at high temperatures, differential solvation due to the increased exposed surface area of the unfolded state and, in particular, its larger thermal volume linked to the increased conformational dynamics of the unfolded state ensemble takes over and ΔV for unfolding eventually becomes positive. 相似文献
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Dr. Alexander R. Nödling Dr. Katarzyna Świderek Dr. Raquel Castillo Jonathan W. Hall Dr. Antonio Angelastro Dr. Louis C. Morrill Dr. Yi Jin Dr. Yu‐Hsuan Tsai Prof. Vicent Moliner Dr. Louis Y. P. Luk 《Angewandte Chemie (International ed. in English)》2018,57(38):12478-12482
There has been growing interest in performing organocatalysis within a supramolecular system as a means of controlling reaction reactivity and stereoselectivity. Here, a protein is used as a host for iminium catalysis. A pyrrolidine moiety is covalently linked to biotin and introduced to the protein host streptavidin for organocatalytic activity. Whereas in traditional systems stereoselectivity is largely controlled by the substituents added to the organocatalyst, enantiomeric enrichment by the reported supramolecular system is completely controlled by the host. Also, the yield of the model reaction increases over 10‐fold when streptavidin is included. A 1.1 Å crystal structure of the protein–catalyst complex and molecular simulations of a key intermediate reveal the chiral scaffold surrounding the organocatalytic reaction site. This work illustrates that proteins can be an excellent supramolecular host for driving stereoselective secondary amine organocatalysis. 相似文献
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Muhammad Ehsan Ashwani Kumar Jonas S. Mortensen Yang Du Parameswaran Hariharan Kaavya K. Kumar Betty Ha Bernadette Byrne Lan Guan Brian K. Kobilka Claus J. Loland Pil Seok Chae 《化学:亚洲杂志》2019,14(11):1926-1931
We prepared an amphiphile with a penta‐phenylene lipophilic group and a branched trimaltoside head group. This new agent, designated penta‐phenylene maltoside (PPM), showed a marked tendency to self‐assembly into micelles via strong aromatic–aromatic interactions in aqueous media, as evidenced by 1H NMR spectroscopy and fluorescence studies. When utilized for membrane protein studies, this new agent was superior to DDM, a gold standard conventional detergent, in stabilizing multiple proteins long term. The ability of this agent to form aromatic–aromatic interactions is likely responsible for enhanced protein stabilization when associated with a target membrane protein. 相似文献
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An Artificial Enzyme Made by Covalent Grafting of an FeII Complex into β‐Lactoglobulin: Molecular Chemistry,Oxidation Catalysis,and Reaction‐Intermediate Monitoring in a Protein 下载免费PDF全文
Charlotte Buron Dr. Katell Sénéchal‐David Dr. Rémy Ricoux Jean‐Pierre Le Caër Vincent Guérineau Dr. Philippe Méjanelle Dr. Régis Guillot Dr. Christian Herrero Prof. Jean‐Pierre Mahy Prof. Frédéric Banse 《Chemistry (Weinheim an der Bergstrasse, Germany)》2015,21(34):12188-12193
An artificial metalloenzyme based on the covalent grafting of a nonheme FeII polyazadentate complex into bovine β‐lactoglobulin has been prepared and characterized by using various spectroscopic techniques. Attachment of the FeII catalyst to the protein scaffold is shown to occur specifically at Cys121. In addition, spectrophotometric titration with cyanide ions based on the spin‐state conversion of the initial high spin (S=2) FeII complex into a low spin (S=0) one allows qualitative and quantitative characterization of the metal center’s first coordination sphere. This biohybrid catalyst activates hydrogen peroxide to oxidize thioanisole into phenylmethylsulfoxide as the sole product with an enantiomeric excess of up to 20 %. Investigation of the reaction between the biohybrid system and H2O2 reveals the generation of a high spin (S=5/2) FeIII(η2‐O2) intermediate, which is proposed to be responsible for the catalytic sulfoxidation of the substrate. 相似文献
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Thomas G. Back Ziad Moussa Masood Parvez 《Angewandte Chemie (Weinheim an der Bergstrasse, Germany)》2004,116(10):1288-1290
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Perczel A Jákli I McAllister MA Csizmadia IG 《Chemistry (Weinheim an der Bergstrasse, Germany)》2003,9(11):2551-2566
Folding properties of small globular proteins are determined by their amino acid sequence (primary structure). This holds both for local (secondary structure) and for global conformational features of linear polypeptides and proteins composed from natural amino acid derivatives. It thus provides the rational basis of structure prediction algorithms. The shortest secondary structure element, the beta-turn, most typically adopts either a type I or a type II form, depending on the amino acid composition. Herein we investigate the sequence-dependent folding stability of both major types of beta-turns using simple dipeptide models (-Xxx-Yyy-). Gas-phase ab initio properties of 16 carefully selected and suitably protected dipeptide models (for example Val-Ser, Ala-Gly, Ser-Ser) were studied. For each backbone fold most probable side-chain conformers were considered. Fully optimized 321G RHF molecular structures were employed in medium level [B3LYP/6-311++G(d,p)//RHF/3-21G] energy calculations to estimate relative populations of the different backbone conformers. Our results show that the preference for beta-turn forms as calculated by quantum mechanics and observed in Xray determined proteins correlates significantly. 相似文献
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The RING Domain of the Scaffold Protein Ste5 Adopts a Molten Globular Character with High Thermal and Chemical Stability 下载免费PDF全文
Michal J. Walczak Dr. Brighton Samatanga Dr. Frank van Drogen Prof. Dr. Matthias Peter Dr. Ilian Jelesarov Prof. Dr. Gerhard Wider 《Angewandte Chemie (International ed. in English)》2014,53(5):1320-1323
Ste5 is a scaffold protein that controls the pheromone response of the MAP‐kinase cascade in yeast cells. Upon pheromone stimulation, Ste5 (through its RING‐H2 domain) interacts with the β and γ subunits of an activated heterodimeric G protein and promotes activation of the MAP‐kinase cascade. With structural and biophysical studies, we show that the Ste5 RING‐H2 domain exists as a molten globule under native buffer conditions, in yeast extracts, and even in denaturing conditions containing urea (7 M ). Furthermore, it exhibits high thermal stability in native conditions. Binding of the Ste5 RING‐H2 domain to the physiological Gβ/γ (Ste4/Ste18) ligand is accompanied by a conformational transition into a better folded, more globular structure. This study reveals novel insights into the folding mechanism and recruitment of binding partners by the Ste5 RING‐H2 domain. We speculate that many RING domains may share a similar mechanism of substrate recognition and molten‐globule‐like character. 相似文献
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《Analytical letters》2012,45(15-16):1653-1678
Abstract The kinetic and thermodynamic properties of the folate interaction with immobilized folate binding protein (FBP) are examined. The use of enzyme-rather than radio-labeled folate provides insight into the complex binding mechanism of folate with FBP and indicates that polymerization of the binding protein (evident for FBP-folate association in solution) is not a prerequisite for the cooperative behavior observed. An enzyme-linked competitive binding assay for folate based on this interaction is described and dose-response curves demonstrate the sensitivity and selectivity of the method. The accuracy of the assay is tested by determining folate in infant formula. 相似文献