1.7.1.13 2-amino-5-cyanopyrrolo[2,3-d]pyrimidin-4-one + 2 NADPH + 2 H+ natural substrate Escherichia coli 2-amino-5-amino-methyl-pyrrolo[2,3-d]pyrimidin-4-one + 2 NADP+ - ? 424479 1.7.1.13 2-amino-5-cyanopyrrolo[2,3-d]pyrimidine + 2 NADPH + 2 H+ analogue of natural substrate, poor substrate Escherichia coli 2-amino-5-amino-methyl-pyrrolo[2,3-d]pyrimidine + 2 NADP+ - ? 425399 1.7.1.13 5-cyanopyrrolo[2,3-d]pyrimidin-4-one + 2 NADPH + 2 H+ analogue of natural substrate Escherichia coli 5-amino-methyl-pyrrolo[2,3-d]pyrimidin-4-one + 2 NADP+ - ? 425798 1.7.1.13 7-cyano-7-carba-2-deaminoguanine + 2 NADPH + 2 H+ - Escherichia coli 7-aminomethyl-7-carba-2-deaminoguanine + 2 NADP+ - ? 440974 1.7.1.13 7-cyano-7-carbaguanine + 2 NADPH + 2 H+ - Escherichia coli 7-aminomethyl-7-carbaguanine + 2 NADP+ - ? 440975 1.7.1.13 7-cyano-7-carbaguanine + 2 NADPH + 2 H+ - Vibrio cholerae O1 7-aminomethyl-7-carbaguanine + 2 NADP+ - ? 440975 1.7.1.13 7-cyano-7-carbaguanine + 2 NADPH + 2 H+ - Pectobacterium carotovorum subsp. carotovorum 7-aminomethyl-7-carbaguanine + 2 NADP+ - ? 440975 1.7.1.13 7-cyano-7-carbaguanine + 2 NADPH + 2 H+ - Vibrio cholerae O1 ATCC 39315 7-aminomethyl-7-carbaguanine + 2 NADP+ - ? 440975 1.7.1.13 7-cyano-7-carbaguanine + 2 NADPH + 2 H+ - Pectobacterium carotovorum subsp. carotovorum PCC21 7-aminomethyl-7-carbaguanine + 2 NADP+ - ? 440975 1.7.1.13 7-cyano-7-deazaguanine + 2 NADPH + 2 H+ - Escherichia coli queuine + 2 NADP+ - ? 378400 1.7.1.13 7-cyano-7-deazaguanine + 2 NADPH + 2 H+ - Vibrio cholerae queuine + 2 NADP+ queuine is 7-aminomethyl-7-deazaguanine ? 378400 1.7.1.13 7-cyano-7-deazaguanine + 2 NADPH + 2 H+ late step in biosynthesis of the modified tRNA nucleoside queuosine Bacillus subtilis queuine + 2 NADP+ - ? 378400 1.7.1.13 7-cyano-7-deazaguanine + 2 NADPH + 2 H+ late step in biosynthesis of the modified tRNA nucleoside queuosine Escherichia coli queuine + 2 NADP+ - ? 378400 1.7.1.13 7-cyano-7-deazaguanine + 2 NADPH + 2 H+ - Vibrio cholerae ATCC 39315 queuine + 2 NADP+ queuine is 7-aminomethyl-7-deazaguanine ? 378400 1.7.1.13 7-cyano-7-deazaguanine + 2 NADPH + 2 H+ i.e. preQ0 Escherichia coli 7-aminomethyl-7-deazaguanine + 2 NADP+ - ? 461499 1.7.1.13 7-cyano-7-deazaguanine + NADPH + H+ - Escherichia coli queuine + NADP+ + H+ - ? 425836 1.7.1.13 additional information no substrates: acetonitrile, benzonitrile and benzylcyanide Escherichia coli ? - ? 89 1.7.1.13 additional information during catalysis each active site of the dimeric enzyme binds one substrate molecule. NADPH binds independent of the substrate. The PreQ0 binding pocket of the active site is not involved in the binding of NADPH Escherichia coli ? - ? 89 1.7.1.13 additional information enzyme is highly specific for substrate preQ0 Pectobacterium carotovorum subsp. carotovorum ? - ? 89 1.7.1.13 additional information QueF binds substrate preQ0 in a strongly exothermic process (DeltaH 80.3 kJ/mol) whereby the thioimide adduct is formed with half-of-the-sites reactivity in the homodimeric enzyme. Both steps of preQ0 reduction involve transfer of the 4-pro-R-hydrogen from NADPH. They proceed about 4-7fold more slowly than trapping of the enzyme-bound preQ0 as covalent thioimide and are mainly rate-limiting for the enzyme's kcat Escherichia coli ? - ? 89 1.7.1.13 additional information the nitrile to amine conversion proceeds through four major stages: formation of a C-S covalent bond between the substrate and the catalytic cysteine residue to form the thioimidate intermediate, hydride transfer from NADPH to the substrate to generate the thiohemiaminal intermediate, cleavage of the C-S covalent bond to generate the imine intermediate, and second hydride transfer from NADPH to the imine intermediate to generate the final amine product. The free energy barrier for the rate-limiting step, i.e. the second hydride transfer, is20.8 kcal/mol Vibrio cholerae O1 ? - ? 89 1.7.1.13 additional information the nitrile to amine conversion proceeds through four major stages: formation of a C-S covalent bond between the substrate and the catalytic cysteine residue to form the thioimidate intermediate, hydride transfer from NADPH to the substrate to generate the thiohemiaminal intermediate, cleavage of the C-S covalent bond to generate the imine intermediate, and second hydride transfer from NADPH to the imine intermediate to generate the final amine product. The free energy barrier for the rate-limiting step, i.e. the second hydride transfer, is20.8 kcal/mol Vibrio cholerae O1 ATCC 39315 ? - ? 89 1.7.1.13 additional information enzyme is highly specific for substrate preQ0 Pectobacterium carotovorum subsp. carotovorum PCC21 ? - ? 89