Inhibitor Discovery of Full-Length New Delhi Metallo-β-Lactamase-1 (NDM-1)

Shen, Bingzheng; Yu, Yan; Chen, Hui; Cao, Xin; Lao, Xingzhen; Fang, Yongliang; Shi, Yun; Chen, Jiao; Zheng, Heng

doi:10.1371/journal.pone.0062955

Cited by 32 publications

(31 citation statements)

References 32 publications

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“…Proteins were expressed in E. coli BL21(DE3) cells and purified by affinity chromatography as described previously (12). The purified protein was dialyzed at 4°C against 50 mM HEPES buffer (pH 7.0) supplemented with 250 mM NaCl and 100 M ZnCl 2 .…”

Section: Methodsmentioning

confidence: 99%

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Role of Non-Active-Site Residue Trp-93 in the Function and Stability of New Delhi Metallo-β-Lactamase 1

Khan

Rehman

2016

Antimicrob Agents Chemother

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New Delhi metallo-␤-lactamase-1 (NDM-1) is expressed by various members of Enterobacteriaceae as a defense mechanism to hydrolyze ␤-lactam antibiotics. Despite various studies showing the significance of active-site residues in the catalytic mechanism, there is a paucity of reports addressing the role of non-active-site residues in the structure and function of NDM-1. In this study, we investigated the significance of non-active-site residue Trp-93 in the structure and function of NDM-1. We cloned bla NDM-1 from an Enterobacter cloacae clinical strain (EC-15) and introduced the mutation of Trp-93 to Ala (yielding the Trp93Ala mutant) by PCR-based site-directed mutagenesis. Proteins were expressed and purified to homogeneity by affinity chromatography. The MICs of the Trp93Ala mutant were reduced 4-to 8-fold for ampicillin, cefotaxime, ceftazidime, cefoxitin, imipenem, and meropenem. The poor hydrolytic activity of the Trp93Ala mutant was also reflected by its reduced catalytic efficiency. The overall catalytic efficiency of the Trp93Ala mutant was reduced by 40 to 55% (the K m was reduced, while the k cat was similar to that of wild-type NDM-1 [wtNDM-1]). Heat-induced denaturation showed that the ⌬G D o and T m of Trp93Ala mutant were reduced by 1.8 kcal/mol and 4.8°C, respectively. Far-UV circular dichroism (CD) analysis showed that the ␣-helical content of the Trp93Ala mutant was reduced by 2.9%. The decrease in stability and catalytic efficiency of the Trp93Ala mutant was due to the loss of two hydrogen bonds with Ser-63 and Val-73 and hydrophobic interactions with Leu-65, Val-73, Gln-123, and Asp-124. The study provided insight into the role of non-active-site amino acid residues in the hydrolytic mechanism of NDM-1.

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Section: Methodsmentioning

confidence: 99%

“…All the experiments were carried out in 50 mM HEPES buffer (pH 7.0) containing 250 mM NaCl and 100 M ZnCl 2 at 30°C as described previously (12). Bovine serum albumin (BSA; 20 g/ml) was added in the dilution buffer to prevent denaturation of NDM-1 and Trp93Ala mutant proteins.…”

Section: Methodsmentioning

confidence: 99%

Role of Non-Active-Site Residue Trp-93 in the Function and Stability of New Delhi Metallo-β-Lactamase 1

Khan

Rehman

2016

Antimicrob Agents Chemother

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“…The expression of VIM‐2 was induced by 0.2 mM IPTG when the absorbance at 600 nm reached 1.0–1.5. The cells were disrupted by sonication, and the supernatant of the cell lysate was further purified by nickel‐chelating affinity method as previous described 43…”

Section: Methodsmentioning

confidence: 99%

Simulation of Cake Filtration for Polydisperse Particles

Becker¹,

Cheng²,

Kronsbein³

et al. 2016

Chem Eng & Technol

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When simulating the buildup of a filter cake consisting of polydisperse particles, the choice of the numerical resolution is difficult as particles may cover a large range of diameters. A method was developed that allows to model cake filtration at any resolution. Resolution-dependent fitting parameters can be found such that the solidity and resistivity of the resulting filter cake will be independent of the chosen resolution. The cake buildup is simulated on the filter media scale. Flow through the porous parts is modeled with the Stokes-Brinkman equation, which requires the local solidity and flow resistivity of each porous grid cell as input. It is described how to set the local solidity and resistivity such that a given global solidity and flow resistivity of the filter cake is achieved and the simulated pressure drop is independent of the chosen resolution.

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“…Using the two-stage protein flexible docking protocol mentioned in the methodology section [43], Shen et al established an interaction model for NDM-1 and the thiophene-carboxylic acid inhibitor. The results revealed that sulfur atoms of ligand might interfere with the water bridge and zinc ions in the active site of NDM-1, which provide useful clues for the rational design of effective NDM-1 inhibitors [78].…”

Section: Cases Of Hit Discoverymentioning

confidence: 99%

Computational methods for drug design and discovery: focus on China

Zheng

Liu

Yue

et al. 2013

Trends in Pharmacological Sciences

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In the past decades, China's computational drug design and discovery research has experienced fast development through various novel methodologies. Application of these methods spans a wide range, from drug target identification to hit discovery and lead optimization. In this review, we firstly provide an overview of China's status in this field and briefly analyze the possible reasons for this rapid advancement. The methodology development is then outlined. For each selected method, a short background precedes an assessment of the method with respect to the needs of drug discovery, and, in particular, work from China is highlighted. Furthermore, several successful applications of these methods are illustrated. Finally, we conclude with a discussion of current major challenges and future directions of the field.

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Inhibitor Discovery of Full-Length New Delhi Metallo-β-Lactamase-1 (NDM-1)

Cited by 32 publications

References 32 publications

Role of Non-Active-Site Residue Trp-93 in the Function and Stability of New Delhi Metallo-β-Lactamase 1

Role of Non-Active-Site Residue Trp-93 in the Function and Stability of New Delhi Metallo-β-Lactamase 1

Simulation of Cake Filtration for Polydisperse Particles

Computational methods for drug design and discovery: focus on China

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