Addition of deoxyribose to guanine and modified DNA based by Lactobacillus helveticus trans-N-deoxyribosylase.
The use of bacterial trans-N-deoxyribosylase was evaluated as an alternative method for deoxyribosylation in the synthesis of deoxyribonucleosides containing potentially mutagenic adducts. A crude enzyme preparation was isolated from Lactobacillus helveticus and compared to Escherichia coli purine nucleoside phosphorylase. trans-N-deoxyribosylase was more regioselective than purine nucleoside phosphorylase in the deoxyribosylation of Gua at the N9 atom, as compared to N7, as demonstrated by NMR analysis of the product. 5,6,7,9-Tetrahydro-7-acetoxy-9-oxoimidazo[1,2-a]purine was efficiently deoxyribosylated by trans-N-deoxyribosylase but not at all by purine nucleoside phosphorylase. Other substrates for trans-N-deoxyribosylase were N2-(2-oxoethyl)Gua, pyrimido[1,2-a]purin-10(3H)-one, 1,N2-epsilon-Gua, N2,3-epsilon-Gua, 3,N4-epsilon-Cyt, 1,N6-epsilon-Ade, C8-methylGua, and C8-aminoGua, most of which gave the desired isomer (bond at the nitrogen corresponding to N9 in Gua) in good yield. Neither N7-alkylpurines nor C8-(arylamino)-substituted guanines were substrates. The approach offers a relatively convenient method of enzymatic preparation of many carcinogen-DNA adducts at the nucleoside level, for either use as standards or incorporation into oligonucleotides. trans-N-deoxyribosylase can also be used to remove deoxyribose from modified deoxyribonucleosides in the presence of excess Cyt.
Department of Biochemistry, Vanderbilt University School of Medicine, Nashville, Tennessee 37232, USA.
M
M
Müller
L K
LK
Hutchinson
F P
FP
Guengerich
D018736:DNA Adducts / Q000187:drug effects* / Q000378:metabolism*; D003855:Deoxyribose / Q000235:genetics / Q000378:metabolism*; D006147:Guanine / Q000378:metabolism*; D007778:Lactobacillus / Q000201:enzymology*; D010430:Pentosyltransferases / Q000494:pharmacology*
D018736:DNA Adducts; D003855:Deoxyribose; D006147:Guanine; D010430:Pentosyltransferases; C021888:nucleoside deoxyribosyltransferase