J Michael Sauder - Discovery and structure determination of the orphan enzyme isoxanthopterin deaminase

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      Publication Details (including relevant citation   information):

      Biochemistry (2010) 49:4373-4382.    


      Two previously uncharacterized proteins have been identified   that  efficiently catalyze the deamination of   isoxanthopterin and pterin  6-carboxylate. The genes   encoding these two enzymes, NYSGXRC-9339a (  gi|44585104 )   and NYSGXRC-9236b ( gi|44611670 ), were first identified    from DNA isolated from the Sargasso Sea as part of the Global   Ocean  Sampling Project. The genes were synthesized, and the   proteins were  subsequently expressed and purified. The   X-ray structure of Sgx9339a was  determined at 2.7 A   resolution (Protein Data Bank entry 2PAJ ). This  protein   folds as a distorted (beta/alpha)(8) barrel and contains a    single zinc ion in the active site. These enzymes are members of   the  amidohydrolase superfamily and belong to cog0402 within   the clusters of  orthologous groups (COG). Enzymes in   cog0402 have previously been shown  to catalyze the   deamination of guanine, cytosine,  S-adenosylhomocysteine,   and 8-oxoguanine. A small compound library of  pteridines,   purines, and pyrimidines was used to probe catalytic    activity. The only substrates identified in this search   were  isoxanthopterin and pterin 6-carboxylate. The kinetic   constants for the  deamination of isoxanthopterin with   Sgx9339a were determined to be 1.0  s(-1), 8.0 muM, and 1.3   x 10(5) M(-1) s(-1) (k(cat), K(m), and  k(cat)/K(m),   respectively). The active site of Sgx9339a most closely    resembles the active site for 8-oxoguanine deaminase (Protein   Data Bank  entry 2UZ9 ). A model for substrate recognition   of isoxanthopterin by  Sgx9339a was proposed on the basis of   the binding of guanine and  xanthine in the active site of   guanine deaminase. Residues critical for  substrate binding   appear to be conserved glutamine and tyrosine residues  that   form hydrogen bonds with the carbonyl oxygen at C4, a   conserved  threonine residue that forms hydrogen bonds with   N5, and another  conserved threonine residue that forms   hydrogen bonds with the carbonyl  group at C7. These   conserved active site residues were used to identify  24   other genes which are predicted to deaminate isoxanthopterin.

      Address (URL): http://pubs.acs.org/doi/abs/10.1021/bi100252s