scholarly journals GMP and IMP Are Competitive Inhibitors of CMY-10, an Extended-Spectrum Class C β-Lactamase

2017 ◽  
Vol 61 (5) ◽  
Author(s):  
Jung-Hyun Na ◽  
Young Jun An ◽  
Sun-Shin Cha
Keyword(s):  
Crystal Structure ◽  
Active Site ◽  
Binding Mode ◽  
Competitive Inhibitor ◽  
Phosphate Group ◽  
The Other ◽  
Extended Spectrum ◽  
Competitive Inhibitors ◽  
Class C

ABSTRACT Nucleotides were effective in inhibiting the class C β-lactamase CMY-10. IMP was the most potent competitive inhibitor, with a Ki value of 16.2 μM. The crystal structure of CMY-10 complexed with GMP or IMP revealed that nucleotides fit into the R2 subsite of the active site with a unique vertical binding mode where the phosphate group at one terminus is deeply bound in the subsite and the base at the other terminus faces the solvent.

scholarly journals Structural modeling of GSK3β implicates the inactive (DFG-out) conformation as the target bound by TDZD analogs

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Meenakshisundaram Balasubramaniam ◽  
Nirjal Mainali ◽  
Suresh Kuarm Bowroju ◽  
Paavan Atluri ◽  
Narsimha Reddy Penthala ◽  
...  
Keyword(s):  
Glycogen Synthase ◽  
Three Dimensional ◽  
Binding Mode ◽  
Competitive Inhibitor ◽  
Dimensional Structure ◽  
High Potency ◽  
Subsequent Formation ◽  
Inactive Conformation ◽  
Competitive Inhibitors ◽  
Physiological Pathways

Abstract Glycogen synthase kinase-3β (GSK3β) controls many physiological pathways, and is implicated in many diseases including Alzheimer’s and several cancers. GSK3β-mediated phosphorylation of target residues in microtubule-associated protein tau (MAPTAU) contributes to MAPTAU hyperphosphorylation and subsequent formation of neurofibrillary tangles. Inhibitors of GSK3β protect against Alzheimer’s disease and are therapeutic for several cancers. A thiadiazolidinone drug, TDZD-8, is a non-ATP-competitive inhibitor targeting GSK3β with demonstrated efficacy against multiple diseases. However, no experimental data or models define the binding mode of TDZD-8 with GSK3β, which chiefly reflects our lack of an established inactive conformation for this protein. Here, we used metadynamic simulation to predict the three-dimensional structure of the inactive conformation of GSK3β. Our model predicts that phosphorylation of GSK3β Serine9 would hasten the DFG-flip to an inactive state. Molecular docking and simulation predict the TDZD-8 binding conformation of GSK3β to be inactive, and are consistent with biochemical evidence for the TDZD-8–interacting residues of GSK3β. We also identified the pharmacophore and assessed binding efficacy of second-generation TDZD analogs (TDZD-10 and Tideglusib) that bind GSK3β as non-ATP-competitive inhibitors. Based on these results, the predicted inactive conformation of GSK3β can facilitate the identification of novel GSK3β inhibitors of high potency and specificity.

scholarly journals Conformational Change Observed in the Active Site of Class C β-Lactamase MOX-1 upon Binding to Aztreonam

2015 ◽  
Vol 59 (8) ◽  
pp. 5069-5072 ◽  
Author(s):  
Takuma Oguri ◽  
Yoshikazu Ishii ◽  
Akiko Shimizu-Ibuka
Keyword(s):  
Crystal Structure ◽  
Conformational Change ◽  
Conformational Changes ◽  
Active Site ◽  
Main Chain ◽  
Amide Nitrogen ◽  
Class C

ABSTRACTWe solved the crystal structure of the class C β-lactamase MOX-1 complexed with the inhibitor aztreonam at 1.9Å resolution. The main-chain oxygen of Ser315 interacts with the amide nitrogen of aztreonam. Surprisingly, compared to that in the structure of free MOX-1, this main-chain carboxyl changes its position significantly upon binding to aztreonam. This result indicates that the interaction between MOX-1 and β-lactams can be accompanied by conformational changes in the B3 β-strand main chain.

scholarly journals Carbonic Anhydrase Activators. Part 191 Spectroscopic and Kinetic Investigations for the Interaction of Isozymes I and II With Primary Amines

1997 ◽  
Vol 4 (4) ◽  
pp. 221-227 ◽  
Author(s):  
Fabrizio Briganti ◽  
Andrea Scozzafava ◽  
Claudiu T. Supuran
Keyword(s):  
Carbonic Anhydrase ◽  
Active Site ◽  
Metal Ion ◽  
Competitive Inhibitor ◽  
The Other ◽  
Primary Amines ◽  
Hydrophobic Pocket ◽  
X Ray ◽  
Active Site Cavity ◽  
Kinetic Investigations

The interactions of Zn(II)- and Co(II)-substituted carbonic anhydrase (CA) isozymes I and II with amine type activators such as histamine, serotonin, phenetylamine dopamine and benzylhydrazine have been investigated kinetically, and spectroscopically. All of such activators are of the non-competitive type towards CO2 hydration and 4-nitrophenylacetate hydrolysis for both human isozymes (HCA I and HCA II). The electronic spectra of the adducts of Co(II)CA with amine activators are similar to the spectrum of the previously reported Co(II)CAII-phenol adduct, the only known competitive inhibitor towards CO2 hydration, where the phenol molecule binds into the hydrophobic pocket of the active site. This is a direct spectroscopic evidence that the activator molecules bind within the active site, but not directly to the metal ion. Recent X-ray crystallographic data for the adduct of HCA II with histamine show that the activator molecule is bound at the entrance of the active site cavity, near to residues His 64, Asn 62 and Gln 92, where actively aids in shuttling protons between the active site and the environment. Similar arrangements probably occur for the other activators reported in the present paper.

scholarly journals Modified Penicillin Molecule with Carbapenem-Like Stereochemistry Specifically Inhibits Class C β-Lactamases

2017 ◽  
Vol 61 (12) ◽  
Author(s):  
Xuehua Pan ◽  
Yunjiao He ◽  
Tianfeng Chen ◽  
Kin-Fai Chan ◽  
Yanxiang Zhao
Keyword(s):  
Active Site ◽  
Binding Mode ◽  
Carboxyl Group ◽  
Hydrogen Atoms ◽  
Chemical Modifications ◽  
Inhibitory Potency ◽  
Current Collection ◽  
Class A ◽  
Lactam Ring ◽  
Class C

ABSTRACT Bacterial β-lactamases readily inactivate most penicillins and cephalosporins by hydrolyzing and “opening” their signature β-lactam ring. In contrast, carbapenems resist hydrolysis by many serine-based class A, C, and D β-lactamases due to their unique stereochemical features. To improve the resistance profile of penicillins, we synthesized a modified penicillin molecule, MPC-1, by “grafting” carbapenem-like stereochemistry onto the penicillin core. Chemical modifications include the trans conformation of hydrogen atoms at C-5 and C-6 instead of cis, and a 6-α hydroxyethyl moiety to replace the original 6-β aminoacyl group. MPC-1 selectively inhibits class C β-lactamases, such as P99, by forming a nonhydrolyzable acyl adduct, and its inhibitory potency is ∼2 to 5 times higher than that for clinically used β-lactamase inhibitors clavulanate and sulbactam. The crystal structure of MPC-1 forming the acyl adduct with P99 reveals a novel binding mode for MPC-1 that resembles carbapenem bound in the active site of class A β-lactamases. Furthermore, in this novel binding mode, the carboxyl group of MPC-1 blocks the deacylation reaction by occluding the critical catalytic water molecule and renders the acyl adduct nonhydrolyzable. Our results suggest that by incorporating carbapenem-like stereochemistry, the current collection of over 100 penicillins and cephalosporins can be modified into candidate compounds for development of novel β-lactamase inhibitors.

scholarly journals Structural basis for regiospecific midazolam oxidation by human cytochrome P450 3A4

2016 ◽  
Vol 114 (3) ◽  
pp. 486-491 ◽  
Author(s):  
Irina F. Sevrioukova ◽  
Thomas L. Poulos
Keyword(s):  
Crystal Structure ◽  
Cytochrome P450 ◽  
Active Site ◽  
Structural Information ◽  
Binding Mode ◽  
Structural Basis ◽  
Cytochrome P450 3A4 ◽  
Human Cytochrome ◽  
Cyp3a4 Substrate

Human cytochrome P450 3A4 (CYP3A4) is a major hepatic and intestinal enzyme that oxidizes more than 60% of administered therapeutics. Knowledge of how CYP3A4 adjusts and reshapes the active site to regioselectively oxidize chemically diverse compounds is critical for better understanding structure–function relations in this important enzyme, improving the outcomes for drug metabolism predictions, and developing pharmaceuticals that have a decreased ability to undergo metabolism and cause detrimental drug–drug interactions. However, there is very limited structural information on CYP3A4–substrate interactions available to date. Despite the vast variety of drugs undergoing metabolism, only the sedative midazolam (MDZ) serves as a marker substrate for the in vivo activity assessment because it is preferentially and regioselectively oxidized by CYP3A4. We solved the 2.7 Å crystal structure of the CYP3A4–MDZ complex, where the drug is well defined and oriented suitably for hydroxylation of the C1 atom, the major site of metabolism. This binding mode requires H-bonding to Ser119 and a dramatic conformational switch in the F–G fragment, which transmits to the adjacent D, E, H, and I helices, resulting in a collapse of the active site cavity and MDZ immobilization. In addition to providing insights on the substrate-triggered active site reshaping (an induced fit), the crystal structure explains the accumulated experimental results, identifies possible effector binding sites, and suggests why MDZ is predominantly metabolized by the CYP3A enzyme subfamily.

Molecular aspects of β-ketoacyl synthase (KAS) catalysis

2000 ◽  
Vol 28 (6) ◽  
pp. 601-607 ◽  
Author(s):  
P. von Wettstein-Knowles ◽  
J. Gotthardt Olsen ◽  
K. Arnvig McGuire ◽  
S. Larsen
Keyword(s):  
Crystal Structure ◽  
Escherichia Coli ◽  
Active Site ◽  
Chalcone Synthase ◽  
Crystal Structure Data ◽  
The Other ◽  
Ketoacyl Synthase ◽  
Chain Length Specificity ◽  
Catalytic Mechanisms ◽  
Molecular Aspects

Crystal structure data for Escherichia coli β-ketoacyl synthase (KAS) I with C10 and C12 fatty acid substrates bound in conjunction with results from mutagenizing residues in the active site leads to a model for catalysis. Differences from and similarities to the other Claisen enzymes carrying out decarboxylations reveal two catalytic mechanisms, one for KAS I and KAS II, the other for KAS III and chalcone synthase. A comparison of the structures of KAS I and KAS II does not reveal the basis of chain-length specificity. The structures of the Arabidopsis thaliana KAS family are compared.

Structural basis for the extended substrate spectrum of AmpC BER and structure-guided discovery of the inhibition activity of citrate against the class C β-lactamases AmpC BER and CMY-10

2016 ◽  
Vol 72 (8) ◽  
pp. 976-985 ◽  
Author(s):  
Jung-Hyun Na ◽  
Sun-Shin Cha
Keyword(s):  
Active Site ◽  
Active Sites ◽  
Competitive Inhibitor ◽  
Structural Basis ◽  
Active Site Residues ◽  
Weakly Bound ◽  
Primary Metabolite ◽  
Operative Strategy ◽  
Class C ◽  
Substrate Spectrum

AmpC BER is an extended substrate spectrum class C β-lactamase with a two-amino-acid insertion in the R2 loop compared with AmpC EC2. The crystal structures of AmpC BER (S64A mutant) and AmpC EC2 were determined. Structural comparison of the two proteins revealed that the insertion increases the conformational flexibility of the R2 loop. Two citrate molecules originating from the crystallization solution were observed in the active site of the S64A mutant. One citrate molecule makes extensive interactions with active-site residues that are highly conserved among class C β-lactamases, whereas the other one is weakly bound. Based on this structural observation, it is demonstrated that citrate, a primary metabolite that is widely used as a food additive, is a competitive inhibitor of two class C β-lactamases (AmpC BER and CMY-10). Consequently, the data indicate enhancement of the flexibility of the R2 loop as an operative strategy for molecular evolution of extended-spectrum class C β-lactamases, and also suggest that the citrate scaffold is recognized by the active sites of class C β-lactamases.

scholarly journals Crystal Structure of the Mycobacterium fortuitum Class A β-Lactamase: Structural Basis for Broad Substrate Specificity

2006 ◽  
Vol 50 (7) ◽  
pp. 2516-2521 ◽  
Author(s):  
Eric Sauvage ◽  
Eveline Fonzé ◽  
Birgit Quinting ◽  
Moreno Galleni ◽  
Jean-Marie Frère ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Broad Spectrum ◽  
Active Site ◽  
Bacterial Resistance ◽  
Specific Activity ◽  
Structural Basis ◽  
Mycobacterium Fortuitum ◽  
Bacterial Strains ◽  
Class A ◽  
Extended Spectrum

ABSTRACT β-Lactamases are the main cause of bacterial resistance to penicillins and cephalosporins. Class A β-lactamases, the largest group of β-lactamases, have been found in many bacterial strains, including mycobacteria, for which no β-lactamase structure has been previously reported. The crystal structure of the class A β-lactamase from Mycobacterium fortuitum (MFO) has been solved at 2.13-Å resolution. The enzyme is a chromosomally encoded broad-spectrum β-lactamase with low specific activity on cefotaxime. Specific features of the active site of the class A β-lactamase from M. fortuitum are consistent with its specificity profile. Arg278 and Ser237 favor cephalosporinase activity and could explain its broad substrate activity. The MFO active site presents similarities with the CTX-M type extended-spectrum β-lactamases but lacks a specific feature of these enzymes, the VNYN motif (residues 103 to 106), which confers on CTX-M-type extended-spectrum β-lactamases a more efficient cefotaximase activity.

A crystallographic study of the oxidation of lysozyme by iodine

1967 ◽  
Vol 167 (1009) ◽  
pp. 435-438 ◽  
Keyword(s):  
Active Site ◽  
The Other ◽  
Enzymic Hydrolysis ◽  
Substrate Modification ◽  
Crystallographic Study ◽  
Competitive Inhibitors ◽  
Plausible Hypothesis ◽  
The One ◽  
Hydrolysis Of ◽  
Tryptophanyl Residue

The crystallographic examinations of the lysozyme molecule, and of the complexes of lysozyme with competitive inhibitors, have resulted in the identification of the active site and in a plausible hypothesis for its activity as described in the previous papers at this meeting (Blake, Mair, North, Phillips & Sarma, p. 365; Blake, Johnson, Mair, North, Phillips & Sarma, p. 378). The active site includes three tryptophanyl residues (62, 63 and 108) which appear to be involved in the binding of the substrate. Modification of some or all of these residues may provide further evidence of their function in the activity of the enzyme (see figure 19 a of Blake et al ., p. 384). Hartdegen & Rupley (1964) showed that the action of small amounts of triiodide produced two modified lysozymes. One product, containing iodine, was enzymically active, while the other which contained no iodine was inactive. Enzymic hydrolysis of the latter product revealed that it had one less tryptophanyl residue than the native enzyme. Hartdegen & Rupley were not able, however, to show which of the six tryptophanyl residues in the lysozyme molecule was the one uniquely reactive to iodine, but they were able to show that it was part of, or near, the active site.

scholarly journals Crystal structure of papain-E64-c complex. Binding diversity of E64-c to papain S2 and S3 subsites

1992 ◽  
Vol 287 (3) ◽  
pp. 797-803 ◽  
Author(s):  
M J Kim ◽  
D Yamamoto ◽  
K Matsumoto ◽  
M Inoue ◽  
T Ishida ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Active Site ◽  
Cysteine Proteinase ◽  
Complex Structure ◽  
Binding Mode ◽  
Accessible Surface Area ◽  
Cysteine Proteinase Inhibitor ◽  
Binding Modes ◽  
Replacement Method ◽  
Solvent Molecules

In order to investigate the binding mode of E64-c (a synthetic cysteine proteinase inhibitor) to papain at the atomic level, the crystal structure of the complex was analysed by X-ray diffraction at 1.9 A (1 A is expressed in SI units as 0.1 nm) resolution. The crystal has a space group P2(1)2(1)2(1) with a = 43.37, b = 102.34 and c = 49.95 A. A total of 21,135 observed reflections were collected from the same crystal, and 14811 unique reflections of up to 1.9 A resolution [Fo > 3 sigma(Fo)] were used for the structure solution and refinement. The papain structure was determined by means of the molecular replacement method, and then the inhibitor was observed on a (2 magnitude of Fo-magnitude of Fc) difference Fourier map. The complex structure was finally refined to R = 19.4% including 207 solvent molecules. Although this complex crystal (Form II) was polymorphous as compared with the previously analysed one (Form I), the binding modes of leucine and isoamylamide moieties of E64-c were significantly different from each other. By the calculation of accessible surface area for each complex atom, these two different binding modes were both shown to be tight enough to prevent the access of solvent molecules to the papain active site. With respect to the E64-c-papain binding mode, molecular-dynamics simulations proposed two kinds of stationary states which were derived from the crystal structures of Forms I and II. One of these, which corresponds to the binding mode simulated from Form I, was essentially the same as that observed in the crystal structure, and the other was somewhat different from the crystal structure of Form II, especially with respect to the binding of the isoamylamide moiety with the papain S subsites. The substrate specificity for the papain active site is discussed on the basis of the present results.

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