Investigating the position of the hairpin loop in New Delhi metallo-β-lactamase, NDM-1, during catalysis and inhibitor binding
| Autor: | Indra D. Sahu, Abraham Jon Moller, Masaki Horitani, David L. Tierney, Michael W. Crowder, Mahesh Aitha |
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| Rok vydání: | 2016 |
| Předmět: |
0301 basic medicine
Chemistry Mutant Electron Molecular Dynamics Simulation 010402 general chemistry 01 natural sciences Biochemistry Catalysis beta-Lactamases Article 0104 chemical sciences Inorganic Chemistry Metal 03 medical and health sciences Molecular dynamics Crystallography 030104 developmental biology visual_art visual_art.visual_art_medium beta-Lactamase Inhibitors Spin label Spectroscopy Beta-Lactamase Inhibitors |
| Zdroj: | Journal of Inorganic Biochemistry. 156:35-39 |
| ISSN: | 0162-0134 |
| Popis: | In an effort to examine the relative position of a hairpin loop in New Delhi metallo-β-lactamase, NDM-1, during catalysis, rapid freeze quench double electron electron resonance (RFQ-DEER) spectroscopy was used. A doubly-labeled mutant of NDM-1, which had one spin label on the invariant loop at position 69 and another label at position 235, was prepared and characterized. The reaction of the doubly spin labeled mutant with chromacef was freeze quenched at 500 μs and 10 ms. DEER results showed that the average distance between labels decreased by 4 Å in the 500 μs quenched sample and by 2 Å in the 10 ms quenched sample, as compared to the distance in the unreacted enzyme, although the peaks corresponding to distance distributions were very broad. DEER spectra with the doubly spin labeled enzyme with two inhibitors show that the distance between the loop residue at position 69 and the spin label at position 235 does not change upon inhibitor binding. This study suggests that the hairpin loop in NDM-1 moves over the metal ion during the catalysis and then moves back to its original position after hydrolysis, which is consistent with a previous hypothesis based on NMR solution studies on a related metallo-β-lactamase. This study also demonstrates that this loop motion occurs in the millisecond time domain. |
| Databáze: | OpenAIRE |
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