Dissection of the DNA Mimicry of the Bacteriophage T7 Ocr Protein using Chemical Modification
| Autor: | Gareth A. Roberts, C. Logan Mackay, Margaret Nutley, Augoustinos S. Stephanou, Andrew R. Thomson, David T. F. Dryden, Alan Cooper, Laurie P. Cooper, David Clarke |
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| Rok vydání: | 2009 |
| Předmět: |
WT
wild type Protein Folding Methyltransferase Stereochemistry Biology medicine.disease_cause Binding Competitive Article Viral Proteins 03 medical and health sciences chemistry.chemical_compound Structural Biology Bacteriophage T7 medicine QD GdmCl guanidinium hydrochloride Carboxylate SAM S-adenosyl-L-methionine Molecular Biology 030304 developmental biology chemistry.chemical_classification 0303 health sciences EDC 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride restriction/modification system ITC isothermal titration calorimetry MALDI-TOF matrix-assisted laser desorption/ionization time of flight Molecular Mimicry 030302 biochemistry & molecular biology DNA mimic Chemical modification DNA Methyltransferases Ocr overcome classical restriction Molecular mimicry Enzyme MS mass spectrometry chemistry Biochemistry FT-ICR Fourier transform ion cyclotron resonance Nucleic Acid Conformation Restriction modification system Protein folding R/M restriction/modification chemical modification Dimerization HOBt hydroxybenzotriazole |
| Zdroj: | Journal of Molecular Biology Stephanou, A S, Roberts, G A, Cooper, L P, Clarke, D J, Thomson, A R, MacKay, C L, Nutley, M, Cooper, A & Dryden, D T F 2009, ' Dissection of the DNA Mimicry of the Bacteriophage T7 Ocr Protein using Chemical Modification ', Journal of Molecular Biology, vol. 391, no. 3, pp. 565-576 . https://doi.org/10.1016/j.jmb.2009.06.020 |
| ISSN: | 0022-2836 |
| Popis: | The homodimeric Ocr (overcome classical restriction) protein of bacteriophage T7 is a molecular mimic of double-stranded DNA and a highly effective competitive inhibitor of the bacterial type I restriction/modification system. The surface of Ocr is replete with acidic residues that mimic the phosphate backbone of DNA. In addition, Ocr also mimics the overall dimensions of a bent 24-bp DNA molecule. In this study, we attempted to delineate these two mechanisms of DNA mimicry by chemically modifying the negative charges on the Ocr surface. Our analysis reveals that removal of about 46% of the carboxylate groups per Ocr monomer results in an approximately 50-fold reduction in binding affinity for a methyltransferase from a model type I restriction/modification system. The reduced affinity between Ocr with this degree of modification and the methyltransferase is comparable with the affinity of DNA for the methyltransferase. Additional modification to remove approximately 86% of the carboxylate groups further reduces its binding affinity, although the modified Ocr still binds to the methyltransferase via a mechanism attributable to the shape mimicry of a bent DNA molecule. Our results show that the electrostatic mimicry of Ocr increases the binding affinity for its target enzyme by up to approximately 800-fold. |
| Databáze: | OpenAIRE |
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