Complete template-directed enzymatic synthesis of a potential antisense DNA containing 42 methylphosphonodiester bonds.
P alpha-Methyl thymidine triphosphate was prepared through the pyrophosphorolysis of P alpha-methyl thymidine diphosphate P beta-diphenyl ester and tested as an alternative substrate for E. coli DNA polymerase 1 (Klenow fragment) using several template-primer systems requiring the formation of 1 to 42 methylphosphono diester bonds. The enzyme catalyzes the incorporation of a P-methyl thymidylic residue with (Sp)-configuration at a single site in a recessive 3'-end as well as at multiple sites along a growing 167 nucleotide long chain. The synthesis of a full length product, containing 42 sites of methylphosphonate incorporation was observed.
Dineva
M A
MA
Laboratory of Biocatalysis, Bulgarian Academy of Sciences, Sofia.
Chakurov
S
S
Bratovanova
E K
EK
Devedjiev
I
I
Petkov
D D
DD
D001483:Base Sequence; D004256:DNA Polymerase I / Q000378:metabolism; D017931:DNA Primers / Q000737:chemistry; D016373:DNA, Antisense / Q000096:biosynthesis* / Q000138:chemical synthesis* / Q000737:chemistry; D004926:Escherichia coli / Q000201:enzymology; D007700:Kinetics; D008969:Molecular Sequence Data; D013942:Thymine Nucleotides / Q000138:chemical synthesis / Q000737:chemistry
D017931:DNA Primers; D016373:DNA, Antisense; D013942:Thymine Nucleotides; D004256:DNA Polymerase I