Structural and Computational Investigations of VIM-7: Insights into the Substrate Specificity of VIM Metallo-β-Lactamases

Borra, Pardha Saradhi; Leiros, Hanna‐Kirsti S.; Ahmad, Rafi; Spencer, James; Leiros, Ingar; Walsh, Timothy R.; Sundsfjord, Arnfinn; Samuelsen, Ørjan

doi:10.1016/j.jmb.2011.05.035

Cited by 34 publications

(81 citation statements)

References 59 publications

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“…In other MBLs, the low Zn2 preference has been related to the presence of Cys221, and the Zn2 site is proposed to be essential for substrate binding (30,31). Oxidation of Cys221 has also been observed in other subclass B1 MBLs (BcII, SPM-1, VIM-7, and VIM-2 [20][21][22][23]). …”

Section: Resultsmentioning

confidence: 99%

“…2). The area and volume of the antibiotic binding, where ceftazidime is docked into VIM-7 (22), is smaller for GIM-1 (GIM-1 chains A and B, GIM-1-2mol chains A and B, and GIM-1-Ox chain A) than for IMP-1 (PDB 1DD6) and VIM-7 (PDB 2Y87) using the CASTp server (29). The availability of multiple structures from this study also highlights the mobility of both loops 1 and 2, suggesting that, despite the narrower active-site groove of GIM-1, a wide variety of substrates still can be accommodated by reorientation of these regions (that may in particular be associated with alternative hydrogen bonding by the Trp228 side chain).…”

Section: Resultsmentioning

confidence: 99%

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Crystal Structures of Pseudomonas aeruginosa GIM-1: Active-Site Plasticity in Metallo-β-Lactamases

Borra

Samuelsen

Spencer

et al. 2013

Antimicrob Agents Chemother

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Metallo-␤-lactamases (MBLs) have rapidly disseminated worldwide among clinically important Gram-negative bacteria and have challenged the therapeutic use of ␤-lactam antibiotics, particularly carbapenems. The bla GIM-1 gene, encoding one such enzyme, was first discovered in a Pseudomonas aeruginosa isolate from 2002 and has more recently been reported in Enterobacteriaceae. Here, we present crystal structures of GIM-1 in the apo-zinc (metal-free), mono-zinc (where Cys221 was found to be oxidized), and di-zinc forms, providing nine independently refined views of the enzyme. GIM-1 is distinguished from related MBLs in possessing a narrower active-site groove defined by aromatic side chains (Trp228 and Tyr233) at positions normally occupied by hydrophilic residues in other MBLs. Our structures reveal considerable flexibility in two loops (loop 1, residues 60 to 66; loop 2, residues 223 to 242) adjacent to the active site, with open and closed conformations defined by alternative hydrogen-bonding patterns involving Trp228. We suggest that this capacity for rearrangement permits GIM-1 to hydrolyze a wide range of ␤-lactams in spite of possessing a more constrained active site. Our results highlight the structural diversity within the MBL enzyme family.

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Crystal Structures of Pseudomonas aeruginosa GIM-1: Active-Site Plasticity in Metallo-β-Lactamases

Borra

Samuelsen

Spencer

et al. 2013

Antimicrob Agents Chemother

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“…VIM-7, identified in Pseudomonas aeruginosa, was first reported in a clinical isolate from the United States of America in 2001 (14) and is the most divergent VIM enzyme, with only 74% and 77% sequence identity to VIM-1 and VIM-2, respectively. We previously determined the enzyme kinetics (15) and the three-dimensional structure of VIM-7 (16). The enzyme kinetics showed VIM-7 to be an efficient penicillinase and carbapenemase, but VIM-7 displayed variable cephalosporinase activity compared to VIM-1 and VIM-2, with lower activity against cephalosporins having bulky and positively charged C-3 substituents (15).…”

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confidence: 99%

“…Still, Asp120 is conserved in both the dizinc B1 and B3 subclasses, which contain zinc in both the Zn1 (trihistidine) and Zn2 (Asp-Cys/His-His) sites, and in the monozinc B2 subclass, which mainly contains zinc in the Zn2 site (17). Asp120 binds to both Zn2 and the bridging hydroxyl ion in, e.g., the VIM-2, VIM-4, and VIM-7 structures (16,19,20). During catalysis, Asp120 probably orients the new hydroxyl ion after catalysis and keeps the correct orientations in the protonation step during cleavage of the C-N bond in the four-member ␤-lactam ring (17,21).…”

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confidence: 99%

“…We therefore decided to make the same mutation in VIM-7 (F218Y) to analyze the effect of additional hydrogen bonds on the structure and enzyme activity despite the low amino acid sequence identity between VIM-7 and IMP-1 of 28%. The mutation was also selected since the structure analysis of VIM-7 suggested that residue 218 was one possible residue determinant implicated in the reduced catalytic efficiency of VIM-7 compared to VIM-2, particularly for cephalosporins with a positively charged cyclic substituent at the C-3 position, like ceftazidime and cefepime (15,16). The hydrogen binding cluster in VIM-2 from Tyr218 OH to Asn70 O, to the invariant Asp84 OD2, and to Arg121 NH1, is absent in VIM-7 due to Phe218 and results in a proposed increased flexibility and reduced activity of VIM-7 (16).…”

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His224 Alters the R2 Drug Binding Site and Phe218 Influences the Catalytic Efficiency of the Metallo-β-Lactamase VIM-7

Leiros

Skagseth

Edvardsen

et al. 2014

Antimicrob Agents Chemother

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b Metallo-␤-lactamases (MBLs) are the causative mechanism for resistance to ␤-lactams, including carbapenems, in many Gramnegative pathogenic bacteria. One important family of MBLs is the Verona integron-encoded MBLs (VIM). In this study, the importance of residues Asp120, Phe218, and His224 in the most divergent VIM variant, VIM-7, was investigated to better understand the roles of these residues in VIM enzymes through mutations, enzyme kinetics, crystal structures, thermostability, and docking experiments. The tVIM-7-D120A mutant with a tobacco etch virus (TEV) cleavage site was enzymatically inactive, and its structure showed the presence of only the Zn1 ion. The mutant was less thermostable, with a melting temperature (T m ) of 48.5°C, compared to 55.3°C for the wild-type tVIM-7. In the F218Y mutant, a hydrogen bonding cluster was established involving residues Asn70, Asp84, and Arg121. The tVIM-7-F218Y mutant had enhanced activity compared to wild-type tVIM-7, and a slightly higher T m (57.1°C) was observed, most likely due to the hydrogen bonding cluster. Furthermore, the introduction of two additional hydrogen bonds adjacent to the active site in the tVIM-7-H224Y mutant gave a higher thermostability (T m , 62.9°C) and increased enzymatic activity compared to those of the wild-type tVIM-7. Docking of ceftazidime in to the active site of tVIM-7, tVIM-7-H224Y, and VIM-7-F218Y revealed that the side-chain conformations of residue 224 and Arg228 in the L3 loop and Tyr67 in the L1 loop all influence possible substrate binding conformations. In conclusion, the residue composition of the L3 loop, as shown with the single H224Y mutation, is important for activity particularly toward the positively charged cephalosporins like cefepime and ceftazidime.

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Mechanism of imipenem resistance in metallo‐β‐lactamases expressing pathogenic bacterial spp. and identification of potential inhibitors: An in silico approach

Kullappan

Ramaiah

2018

J of Cellular Biochemistry

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The World Health Organization reports that millions of people around the world are infected with antibiotic-resistant bacteria. Such resistance is more common in Pseudomonas aeruginosa, Acinetobacter baumannii, and Klebsiella pneumoniae strains because of the expression of the metallo-β-lactamases (MBLs) namely Imipenemase (IMP)-1, IMP-2, New Delhi metallo-β-lactamases-, Verona imipenemase (VIM)-4, VIM-5, and VIM-7. We did an in silico analysis to understand the resistance mechanism of imipenem at the structural level. Our modeling studies reveal that the VIM-4-imipenem complex has highest binding energy and forms a stable complex as indicated by a consensus score (C-score) value of 5.44. The intense interaction between the substrate and the β-lactamases leads to the increased hydrolysis of the substrate resulting in rapid hydrolysis of the antibiotic imipenem by VIM-4. Virtual screening of compounds from the ZINC database targeting VIM-4 was done, and we found compound ZINC44608383 as the high binding energy compound with the C-score value of 5.58. This compound could be exploited for inhibitor design and development. The current study helps us to understand the resistance mechanism of imipenem in MBL-expressing strains. Also, we have identified a probable inhibitor for VIM-4. We believe that our results will be useful for researchers in designing potent inhibitors for VIM-4.

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Structural and Computational Investigations of VIM-7: Insights into the Substrate Specificity of VIM Metallo-β-Lactamases

Cited by 34 publications

References 59 publications

Crystal Structures of Pseudomonas aeruginosa GIM-1: Active-Site Plasticity in Metallo-β-Lactamases

Crystal Structures of Pseudomonas aeruginosa GIM-1: Active-Site Plasticity in Metallo-β-Lactamases

His224 Alters the R2 Drug Binding Site and Phe218 Influences the Catalytic Efficiency of the Metallo-β-Lactamase VIM-7

Mechanism of imipenem resistance in metallo‐β‐lactamases expressing pathogenic bacterial spp. and identification of potential inhibitors: An in silico approach

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