Dam methylase fromEscherichia coli: kinetic studies using modified DNA oligomers: hemimethylated substrates
| Autor: | Stéphane Marzabal, Ramon Eritja, Vera Thielking, Stephen DuBois, W. Guschlbauer, Angeles Cano |
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| Rok vydání: | 1995 |
| Předmět: |
Site-Specific DNA-Methyltransferase (Adenine-Specific)
Guanine Stereochemistry Molecular Sequence Data 2-Aminopurine Biology Nucleic Acid Denaturation Methylation chemistry.chemical_compound Polydeoxyribonucleotides Dam methylase Escherichia coli Genetics Uracil Hypoxanthine Base Sequence Escherichia coli Proteins DNA Thymine Kinetics chemistry Biochemistry Hypoxanthines DNA methylation Thermodynamics |
| Zdroj: | Digital.CSIC. Repositorio Institucional del CSIC instname |
| ISSN: | 1362-4962 0305-1048 |
| DOI: | 10.1093/nar/23.18.3648 |
| Popis: | 8 pages, 6 figuras, 2 tables.-- PMID: 7478992 [PubMed].-- PMCID: PMC307261. We have measured steady-state kinetics of the N6-adenine methyltransferase Dam Mtase using as substrates non-selfcomplementary tetradecamer duplexs (d[GCCGGATCTAGACG]-d[CGTCTAGATCC-GGC]) containing the hemimethylated GATC target sequence in one or the other strand and modifications in the GATC target sequence of the complementary strands. Modifications included substitution of guanine by hypoxanthine (I), thymine by uracil (U) or 5-ethyl-uracil (E) and adenine by 2,6-diamino-purine (D). Thermodynamic parameters were obtained from the concentration dependence of the melting temperature (Tm) of the duplexes. Large differences in DNA methylation of duplexes containing single dI for dG substitution of the Dam recognition site were observed compared with the canonical substrate, if the substitution involved the top strand (on the G.C rich side). Substitution in either strand by uracil (dU) or 5-ethyluracil (dE) resulted in small perturbation of the methylation patterns. When 2,6-diamino-purine (dD) replaced the adenine to be methylated, small, but significant methylation was observed. The kinetic parameters of the methylation reaction were compared with the thermodynamic free energies and significant correlation was observed. This work has received the support by grants from the European Community (N° SC*-CT90-0472, TSTS) and from the French-Spanish exchange programme 'Picasso'. |
| Databáze: | OpenAIRE |
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