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Conformational Changes in a Macrolide Antibiotic Binding Protein From Mycobacterium smegmatis Upon ADP Binding.

ABSTRACT: Rv3197 (MABP-1), a non-canonical ABC protein in Mycobacterium tuberculosis, has ATPase activity and confers inducible resistance to the macrolide family of antibiotics. Here we have shown that MSMEG_1954, the homolog of Rv3197 in M. smegmatis, has a similar function of conferring macrolide resistance. Crystal structures of apo-MSMEG_1954 (form1 and form 2) and MSMEG_1954 in complex with ADP have been determined. These three structures show that MSMEG_1954 has at least two different conformations we identify as closed state (MSMEG_1954-form 1) and open state (MSMEG_1954-form 2 and MSMEG_1954-ADP). Structural superimposition shows that the MSMEG_1954-form 2 and MSMEG_1954-ADP complex have similar conformation to that observed for MABP-1 and MABP-1-erythromicin complex structure. However, the antibiotic binding pocket in MSMEG_1954-form 1 is completely blocked by the N-terminal accessory domain. When bound by ADP, the N-terminal accessory domain undergoes conformational change, which results in the open of the antibiotic binding pocket. Because of the degradation of N terminal accessory domain in MSMSG_1954-form 2, it is likely to represent a transitional state between MSMEG_1954-form 1 and MSMEG_1954-ADP complex structure.

SUBMITTER: Zhang Q

PROVIDER: S-EPMC8696161 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Protein-protein interactions represent difficult but increasingly important targets for the design of therapeutic compounds able to interfere with biological processes. Recently, fragment-based strategies have been proposed as attractive approaches for the elaboration of protein-protein surface inhibitors from fragment-like molecules. One major challenge in targeting protein-protein interactions is related to the structural adaptation of the protein surface upon molecular recognition. Methods capable of identifying subtle conformational changes of proteins upon fragment binding are therefore required at the early steps of the drug design process. In this report we present a fast NMR method able to probe subtle conformational changes upon fragment binding. The approach relies on the comparison of experimental fragment-induced Chemical Shift Perturbation (CSP) of amine protons to CSP simulated for a set of docked fragment poses, considering the ring-current effect from fragment binding. We illustrate the method by the retrospective analysis of the complex between the anti-apoptotic Bcl-xL protein and the fragment 4'-fluoro-[1,1'-biphenyl]-4-carboxylic acid that was previously shown to bind one of the Bcl-xL hot spots. The CSP-based approach shows that the protein undergoes a subtle conformational rearrangement upon interaction, for residues located in helices [Formula: see text]2, [Formula: see text]3 and the very beginning of [Formula: see text]5. Our observations are corroborated by residual dipolar coupling measurements performed on the free and fragment-bound forms of the Bcl-xL protein. These NMR-based results are in total agreement with previous molecular dynamic calculations that evidenced a high flexibility of Bcl-xL around the binding site. Here we show that CSP of protein amine protons are useful and reliable structural probes. Therefore, we propose to use CSP simulation to assess protein conformational changes upon ligand binding in the fragment-based drug design approach.

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Conformational Changes in a Macrolide Antibiotic Binding Protein From Mycobacterium smegmatis Upon ADP Binding.

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