Polymerase-catalysed incorporation of glucose nucleotides into a DNA duplex.
The enzymatic recognition of six-membered ring nucleoside triphosphates--in particular the 6'-triphosphates of (beta-D-glucopyranosyl)thymine, (2',3'-dideoxy-beta-D-glucopyranosyl)thymine, (3',4'-dideoxy-beta-D-glucopyranosyl)thymine and (2',3'-dideoxy-beta-D-glucopyranosyl)adenine--was investigated. Despite the facts that the pyranose nucleic acids obtained by polymerisation of these monomers do not hybridise in solution with DNA and that the geometry of a DNA strand in a natural duplex differs from that of a pyranose nucleic acid, elongation of the DNA duplex with all four nucleotide analogues by Vent (exo(-)) polymerase was observed. Modelling experiments showed that hydrogen bonds are formed when 2',3'-dideoxy-beta-homo-T building blocks or beta-D-gluco-T building blocks are incorporated opposite adenosine residues in the template but not when they are incorporated opposite thymine residues in the template. The model shows a near perfect alignment of a secondary hydroxy group at the end of the primer and the alpha-phosphate group of the incoming triphosphate. The results of these experiments provide new information on the role of the active site of the enzyme in the polymerisation reaction.
Renders
Marleen
M
Laboratory of Medicinal Chemistry, Rega Institute for Medical Research, Katholieke Universiteit Leuven, Minderbroedersstraat 10, 3000 Leuven, Belgium.
Abramov
Mikhail
M
Froeyen
Matheus
M
Herdewijn
Piet
P
D020134:Catalytic Domain; D004247:DNA / Q000737:chemistry / Q000378:metabolism*; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism*; D005947:Glucose / Q000737:chemistry / Q000378:metabolism*; D008958:Models, Molecular; D015394:Molecular Structure; D009711:Nucleotides / Q000737:chemistry / Q000378:metabolism*
D009711:Nucleotides; D004247:DNA; D004259:DNA-Directed DNA Polymerase; D005947:Glucose