Molecular dynamics comparison of E. coli WrbA apoprotein and holoprotein
Language English Country Germany Media print-electronic
Document type Comparative Study, Journal Article, Research Support, Non-U.S. Gov't
- MeSH
- Apoproteins chemistry isolation & purification metabolism MeSH
- Flavin Mononucleotide chemistry metabolism MeSH
- Spectrometry, Mass, Electrospray Ionization MeSH
- Kinetics MeSH
- Protein Conformation MeSH
- Methionine analogs & derivatives chemistry metabolism MeSH
- Oxidation-Reduction MeSH
- Escherichia coli Proteins chemistry isolation & purification metabolism MeSH
- Repressor Proteins chemistry isolation & purification metabolism MeSH
- Molecular Dynamics Simulation * MeSH
- Protein Stability MeSH
- Protein Binding MeSH
- Binding Sites MeSH
- Structure-Activity Relationship MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
- Comparative Study MeSH
- Names of Substances
- Apoproteins MeSH
- Flavin Mononucleotide MeSH
- Methionine MeSH
- methionine sulfoxide MeSH Browser
- Escherichia coli Proteins MeSH
- Repressor Proteins MeSH
- WrbA protein, E coli MeSH Browser
WrbA is a novel multimeric flavodoxin-like protein of unknown function. A recent high-resolution X-ray crystal structure of E. coli WrbA holoprotein revealed a methionine sulfoxide residue with full occupancy in the FMN-binding site, a finding that was confirmed by mass spectrometry. In an effort to evaluate whether methionine sulfoxide may have a role in WrbA function, the present analyses were undertaken using molecular dynamics simulations in combination with further mass spectrometry of the protein. Methionine sulfoxide formation upon reconstitution of purified apoWrbA with oxidized FMN is fast as judged by kinetic mass spectrometry, being complete in ∼5 h and resulting in complete conversion at the active-site methionine with minor extents of conversion at heterogeneous second sites. Analysis of methionine oxidation states during purification of holoWrbA from bacterial cells reveals that methionine is not oxidized prior to reconstitution, indicating that methionine sulfoxide is unlikely to be relevant to the function of WrbA in vivo. Although the simulation results, the first reported for WrbA, led to no hypotheses about the role of methionine sulfoxide that could be tested experimentally, they elucidated the origins of the two major differences between apo- and holoWrbA crystal structures, an alteration of inter-subunit distance and a rotational shift within the tetrameric assembly.
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