Enzymatic synthesis of DNA on glycerol nucleic acid templates without stable duplex formation between product and template.
Glycerol nucleic acid (GNA) is an interesting alternative base-pairing system based on an acyclic, glycerol-phosphate backbone repeat unit. The question of whether DNA polymerases can catalyze efficient template-dependent synthesis using GNA as the template is of particular interest because GNA is unable to form a stable duplex with DNA. In the present study, we screened a variety of DNA polymerases for GNA-dependent DNA synthesis. We find that Bst DNA polymerase can catalyze full-length DNA synthesis on a dodecamer GNA template. The efficiency of DNA synthesis is increased by replacing adenine with diaminopurine in both the GNA template and the DNA monomers and by the presence of manganese ions. We suggest that the BstDNA polymerase maintains a short, transient region of base-pairing between the DNA product strand and the GNA template, but that stable duplex formation between product and template strands is not required for template-dependent polymerization.
Tsai
Ching-Hsuan
CH
Howard Hughes Medical Institute, Center for Computational and Integrative Biology, Department of Molecular Biology, and Simches Research Center, Massachusetts General Hospital, 185 Cambridge Street, Boston, MA 02144, USA.
Chen
Jingyang
J
Szostak
Jack W
JW
D020029:Base Pairing; D001483:Base Sequence; D002021:Buffers; D002384:Catalysis; D002413:Cations, Divalent / Q000737:chemistry; D002942:Circular Dichroism; D004247:DNA / Q000096:biosynthesis*; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism*; D005990:Glycerol / Q000737:chemistry*; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008345:Manganese / Q000737:chemistry; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D015394:Molecular Structure; D009691:Nucleic Acid Denaturation / Q000187:drug effects; D009696:Nucleic Acids / Q000138:chemical synthesis / Q000737:chemistry*; D009841:Oligonucleotides / Q000138:chemical synthesis / Q000494:pharmacology; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013698:Templates, Genetic