title: Mechanisms of Protein Translocation on DNA Are Differentially Responsive to Water Activity.
abstract: Water plays important but poorly understood roles in the functions of most biomolecules. We are interested in understanding how proteins use diverse search mechanisms to locate specific sites on DNA; here we present a study of the role of closely associated waters in diverse translocation mechanisms. The bacterial DNA adenine methyltransferase, Dam, moves across large segments of DNA using an intersegmental hopping mechanism, relying in part on movement through bulk water. In contrast, other proteins, such as the bacterial restriction endonuclease EcoRI, rely on a sliding mechanism, requiring the protein to stay closely associated with DNA. Here we probed how these two mechanistically distinct proteins respond to well-characterized osmolytes, dimethyl sulfoxide (DMSO), and glycerol. The ability of Dam to move over large segments of DNA is not impacted by either osmolyte, consistent with its minimal reliance on a sliding mechanism. In contrast, EcoRI endonuclease translocation is significantly enhanced by DMSO and inhibited by glycerol, providing further corroboration that these proteins rely on distinct translocation mechanisms. The well-established similar effects of these osmolytes on bulk water, and their differential effects on macromolecule-associated waters, support our results and provide further evidence of the importance of water in interactions between macromolecules and their ligands.
authors.affiliation: Department of Chemistry and Biochemistry, University of California , Santa Barbara, California 93106, United States.
authors.forename: Brigitte S
authors.identifier: http://orcid.org/0000-0001-6233-3319
authors.initials: BS
authors.lastname: Naughton
authors.affiliation: Department of Chemistry and Biochemistry, University of California , Santa Barbara, California 93106, United States.
authors.forename: Norbert O
authors.identifier: http://orcid.org/0000-0001-6032-2704
authors.initials: NO
authors.lastname: Reich
mesh_terms: D001665:Binding Sites; D003451:Cryoprotective Agents / Q000494:pharmacology; D019175:DNA Methylation; D004269:DNA, Bacterial / Q000737:chemistry / Q000378:metabolism*; D015246:Deoxyribonuclease EcoRI / Q000737:chemistry / Q000378:metabolism*; D004121:Dimethyl Sulfoxide / Q000494:pharmacology; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000378:metabolism*; D005990:Glycerol / Q000494:pharmacology; D009995:Osmosis / Q000502:physiology*; D021381:Protein Transport / Q000187:drug effects*; D015265:Site-Specific DNA-Methyltransferase (Adenine-Specific) / Q000737:chemistry / Q000378:metabolism*; D013379:Substrate Specificity; D014867:Water / Q000494:pharmacology*
chemical_list: D003451:Cryoprotective Agents; D004269:DNA, Bacterial; D029968:Escherichia coli Proteins; D014867:Water; D015265:Site-Specific DNA-Methyltransferase (Adenine-Specific); C499294:dam protein, E coli; D015246:Deoxyribonuclease EcoRI; D005990:Glycerol; D004121:Dimethyl Sulfoxide