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Investigation of Possible Inhibitors Against Multidrug‐Resistant New Delhi Metallo‐Beta Lactamase (NDM‐1)
Author(s) -
Hernandez Sonia Lisseth,
Shrestha Sabi,
Beckhem Josh,
Fast Walter L
Publication year - 2017
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.31.1_supplement.921.10
Subject(s) - klebsiella pneumoniae , antimicrobial , virtual screening , antibiotic resistance , bacteria , antibiotics , microbiology and biotechnology , fast protein liquid chromatography , chemistry , enzyme , ion chromatography , biochemistry , biology , drug discovery , escherichia coli , genetics , gene
Since penicillin was introduced in the world of medicine, this antibiotic saved millions from infectious diseases. However, in the past decades, antimicrobial resistance has developed into a widespread public health threat causing high morbidity and mortality rates. New Delhi Metallo β‐lactamase (NDM‐1) is associated with many resistant nosocomial infections by organisms such as Klebsiella pneumoniae . This organism has become resistant by producing NDM‐1 which hydrolyzes the beta‐lactam ring found in beta‐lactam antibiotics. It is important to find novel inhibitors against New Delhi Metallo β‐lactamase (NDM‐1). In order to find potential small molecule inhibitors against NDM‐1, a combination of wet lab and virtual screening procedures were performed. Virtual screening was done to identify high scoring compounds using GOLD. Different virtual libraries were screened along with positive and negative controls. In the wet lab, competent BL21(DE3) bacteria cells were transformed with an expression plasmid containing the CDS of NDM‐1 and then expressed using IPTG and ZnSO 4 to improve soluble yield. In addition, three methods of purification were performed which included nickel affinity chromatography with 6xHIS tag, ion exchange and size exclusion Fast Protein Liquid Chromatography (FPLC). The protein samples were characterized using SDS‐PAGE gel. Furthermore, protein activity of the enzyme was analyzed in the presence of potential inhibitors using Nitrocefin as a substrate in spectrophotometric assays. Differential Scanning Fluorimetry (DSF) was performed to analyze the binding impact of several potential inhibitors on the melting temperature of NDM‐1. Out of the 12 compounds, only 1 showed modest shifts to the melting curve. Any novel compounds found to inhibit NDM‐1 activity may lead to effective new inhibitors to overcome antibiotic resistance.