The Kinetic and Molecular Basis for the Interaction of LexA and Activated RecA Revealed by a Fluorescent Amino Acid Probe
| Autor: | Rahul M. Kohli, Chloe M. Jones, Zachary M Hostetler, Michael B. Cory, E. James Petersson |
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| Jazyk: | angličtina |
| Rok vydání: | 2020 |
| Předmět: |
0301 basic medicine
Models Molecular DNA damage Protein Conformation viruses Repressor Plasma protein binding 01 natural sciences Biochemistry Article 03 medical and health sciences Protein structure Bacterial Proteins Escherichia coli Binding site SOS response Amino Acids Derepression Fluorescent Dyes Binding Sites 010405 organic chemistry Chemistry Escherichia coli Proteins Serine Endopeptidases Drug Resistance Microbial General Medicine biochemical phenomena metabolism and nutrition 0104 chemical sciences Anti-Bacterial Agents DNA-Binding Proteins enzymes and coenzymes (carbohydrates) Kinetics Rec A Recombinases 030104 developmental biology Biophysics Molecular Medicine bacteria Repressor lexA DNA Damage Protein Binding |
| Zdroj: | ACS Chem Biol |
| Popis: | The bacterial DNA damage response (the SOS response) is a key pathway involved in antibiotic evasion and a promising target for combating acquired antibiotic resistance. Activation of the SOS response is controlled by two proteins: the repressor LexA and the DNA damage sensor RecA. Following DNA damage, direct interaction between RecA and LexA leads to de-repression of the SOS response. However, the exact molecular details of this interaction remain unknown. Here, we employ the fluorescent unnatural amino acid acridonylalanine (Acd) as a minimally-perturbing probe of the E. coli RecA:LexA complex. Using LexA labeled with Acd, we report the first kinetic model for the reversible binding of LexA to activated RecA. We also characterize the effects that specific amino acid truncations or substitutions in LexA have on RecA:LexA binding strength, and demonstrate that a mobile loop encoding LexA residues 75‒84 comprises a key recognition interface for RecA. Beyond insights into SOS activation, our approach also further establishes Acd as a sensitive fluorescent probe for investigating the dynamics of protein-protein interactions in other complex systems. |
| Databáze: | OpenAIRE |
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