scholarly journals Structural and biochemical characterization of the nucleoside hydrolase fromC. elegansreveals the role of two active site cysteine residues in catalysis

2017 ◽  
Vol 26 (5) ◽  
pp. 985-996 ◽  
Author(s):  
Ranjan Kumar Singh ◽  
Jan Steyaert ◽  
Wim Versées
Keyword(s):  
Active Site ◽  
Biochemical Characterization ◽  
Cysteine Residues ◽  
Active Site Cysteine ◽  
Nucleoside Hydrolase

Characterization of anEscherichia coliSulfite Oxidase Homologue Reveals the Role of a Conserved Active Site Cysteine in Assembly and Function†

Biochemistry ◽  
2005 ◽  
Vol 44 (30) ◽  
pp. 10339-10348 ◽  
Author(s):  
Stephen J. Brokx ◽  
Richard A. Rothery ◽  
Guijin Zhang ◽  
Derek P. Ng ◽  
Joel H. Weiner
Keyword(s):  
Active Site ◽  
Active Site Cysteine ◽  
And Function

Computational investigations on the catalytic mechanism of maleate isomerase: the role of the active site cysteine residues

2014 ◽  
Vol 16 (24) ◽  
pp. 12462-12474 ◽  
Author(s):  
Hisham M. Dokainish ◽  
Bogdan F. Ion ◽  
James W. Gauld
Keyword(s):  
Active Site ◽  
Catalytic Mechanism ◽  
The State ◽  
Cysteine Residues ◽  
Active Site Cysteine ◽  
Computational Enzymology

Multiscale computational enzymology provides key insights into the state of the substrate-bound active site and roles of its cysteinyl residues.

scholarly journals Structural and biochemical characterization of the environmental MBLs MYO-1, ECV-1 and SHD-1

2020 ◽  
Vol 75 (9) ◽  
pp. 2554-2563 ◽  
Author(s):  
Christopher Fröhlich ◽  
Vidar Sørum ◽  
Sandra Huber ◽  
Ørjan Samuelsen ◽  
Fanny Berglund ◽  
...  
Keyword(s):  
Biochemical Characterization ◽  
Homology Modelling ◽  
Catalytic Efficiency ◽  
Hydrolytic Activity ◽  
Human Pathogens ◽  
E Coli ◽  
X Ray Crystallography ◽  
Environmental Reservoir

Abstract Background MBLs form a large and heterogeneous group of bacterial enzymes conferring resistance to β-lactam antibiotics, including carbapenems. A large environmental reservoir of MBLs has been identified, which can act as a source for transfer into human pathogens. Therefore, structural investigation of environmental and clinically rare MBLs can give new insights into structure–activity relationships to explore the role of catalytic and second shell residues, which are under selective pressure. Objectives To investigate the structure and activity of the environmental subclass B1 MBLs MYO-1, SHD-1 and ECV-1. Methods The respective genes of these MBLs were cloned into vectors and expressed in Escherichia coli. Purified enzymes were characterized with respect to their catalytic efficiency (kcat/Km). The enzymatic activities and MICs were determined for a panel of different β-lactams, including penicillins, cephalosporins and carbapenems. Thermostability was measured and structures were solved using X-ray crystallography (MYO-1 and ECV-1) or generated by homology modelling (SHD-1). Results Expression of the environmental MBLs in E. coli resulted in the characteristic MBL profile, not affecting aztreonam susceptibility and decreasing susceptibility to carbapenems, cephalosporins and penicillins. The purified enzymes showed variable catalytic activity in the order of <5% to ∼70% compared with the clinically widespread NDM-1. The thermostability of ECV-1 and SHD-1 was up to 8°C higher than that of MYO-1 and NDM-1. Using solved structures and molecular modelling, we identified differences in their second shell composition, possibly responsible for their relatively low hydrolytic activity. Conclusions These results show the importance of environmental species acting as reservoirs for MBL-encoding genes.

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