RESUMEN
T:G mismatches in mammals arise primarily from the deamination of methylated CpG sites or the incorporation of improper nucleotides. The process by which repair enzymes such as thymine DNA glycosylase (TDG) identify a canonical DNA base in the incorrect pairing context remains a mystery. However, the abundant contacts of the repair enzymes with the DNA backbone suggest a role for protein-phosphate interaction in the recognition and repair processes, where conformational properties may facilitate the proper interactions. We have previously used 31P NMR to investigate the energetics of DNA backbone BI-BII interconversion and the effect of a mismatch or lesion compared to canonical DNA and found stepwise differences in ΔG of 1-2 kcal/mol greater than equivalent steps in unmodified DNA. We have currently compared our results to substrate dependence for TDG, MBD4, M. HhaI, and CEBPß, testing for correlations to sequence and base-pair dependence. We found strong correlations of our DNA phosphate backbone equilibrium (Keq) to different enzyme kinetics or binding parameters of these varied enzymes, suggesting that the backbone equilibrium may play an important role in mismatch recognition and/or conformational rearrangement and energetics during nucleotide flipping or other aspects of enzyme interrogation of the DNA substrate.
Asunto(s)
Nucleótidos , Timina ADN Glicosilasa , Animales , Conformación Molecular , Nucleótidos/metabolismo , ADN/química , Secuencia de Bases , Timina ADN Glicosilasa/química , Reparación del ADN , Mamíferos/metabolismoRESUMEN
In the main fraction of oligosaccharides obtained after the acetic acid hydrolysis of the lipopolysaccharides of the strain PM were identified: terminal bound glucose and L-glycero-D-manno-heptose, 1,2-,1,3,4-, and 1,3,4,6,-linked heptose. The oligosaccharide of the strain 1297 S contains terminal linked galactose, 1,4- or 1,6-linked glucose and 1,4-linked N-acetylglucosamine additionally. The molar ratios of these elements show the heterogenicity of the preparations. The determined molecular weights underline the R-character of the LPS. In the by-fractions of the acetic acid hydrolysis the known mannan and another polysaccharide with galactose and glucosamine were identified.