7-Cyano-7-deazaguanine synthase (E.C. 6.3.4.20) is an enzyme that catalyzes the formation of a nitrile from a carboxylic acid and ammonia at the expense of ATP. The protein from G. kaustophilus was heterologously expressed, and its biochemical characteristics were explored by using a newly developed HPLC-MS based assay, (31) P NMR, and a fluorescence-based thermal-shift assay. The protein showed the expected high thermostability, had a pH optimum at pH 9.5, and an apparent temperature optimum at 60 °C. We observed strict substrate specificity of QueC for the natural substrate 7-carboxy-7-deazaguanine, and determined AMP and pyrophosphate as co-products of preQ0.
Keywords: 7-cyano-7-deazaguanine synthase; AMP forming; deazapurine; enzyme catalysis; nitrile formation; protein structures.
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