Random Mutagenesis Analysis of the Influenza A M2 Proton Channel Reveals Novel Resistance Mutants
| Autor: | Paul Santner, Jonas S. Laursen, Caroline Kampmeyer, Christian A. Olsen, João M. Martins, Jakob R. Winther, Isaiah T. Arkin, Martin Willemoës, Amelie Stein, Kresten Lindorff-Larsen, Rasmus Hartmann-Petersen |
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| Rok vydání: | 2018 |
| Předmět: |
0301 basic medicine
Drug media_common.quotation_subject Mutant Mutation Missense Mutagenesis (molecular biology technique) Drug resistance 010402 general chemistry Antiviral Agents 01 natural sciences Biochemistry Ion Channels Influenza A Virus H2N2 Subtype Viral Matrix Proteins 03 medical and health sciences Escherichia Drug Resistance Viral Escherichia coli Humans media_common Genetics biology Influenza treatment Chemistry Influenza A Virus H3N2 Subtype biology.organism_classification 0104 chemical sciences 030104 developmental biology Amino Acid Substitution M2 proton channel Mutagenesis biology.protein Function (biology) |
| Zdroj: | Biochemistry. 57:5957-5968 |
| ISSN: | 1520-4995 0006-2960 |
| Popis: | The influenza M2 proton channel is a major drug target, but unfortunately, the acquisition of resistance mutations greatly reduces the functional life span of a drug in influenza treatment. New M2 inhibitors that inhibit mutant M2 channels otherwise resistant to the early adamantine-based drugs have been reported, but it remains unclear whether and how easy resistance could arise to such inhibitors. We have combined a newly developed proton conduction assay with an established method for selection and screening, both Escherichia coli-based, to enable the study of M2 function and inhibition. Combining this platform with two groups of structurally different M2 inhibitors allowed us to isolate drug resistant M2 channels from a mutant library. Two groups of M2 variants emerged from this analysis. A first group appeared almost unaffected by the inhibitor, M_089 (N13I, I35L, and F47L) and M_272 (G16C and D44H), and the single-substitution variants derived from these (I35L, L43P, D44H, and L46P). Functionally, these resemble the known drug resistant M2 channels V27A, S31N, and swine flu. In addition, a second group of tested M2 variants were all still inhibited by drugs but to a lesser extent than wild type M2. Molecular dynamics simulations aided in distinguishing the two groups where drug binding to the wild type and the less resistant M2 group showed a stable positioning of the ligand in the canonical binding pose, as opposed to the drug resistant group in which the ligand rapidly dissociated from the complex during the simulations. |
| Databáze: | OpenAIRE |
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