Escherichia coli 6-pyruvoyltetrahydropterin synthase ortholog encoded by ygcM has a new catalytic activity for conversion of sepiapterin to 7,8-dihydropterin.

The putative gene (ygcM) of Escherichia coli was verified in vitro to encode the ortholog of 6-pyruvoyltetrahydropterin synthase (PTPS). Unexpectedly, the enzyme was found to convert sepiapterin to 7,8-dihydropterin without any cofactors. The enzymatic product 7,8-dihydropterin was identified by HPLC and mass spectrometry analyses, suggesting a novel activity of the enzyme to cleave the C6 side chain of sepiapterin. The optimal activity occurred at pH 6.5-7.0. The reaction rate increased up to 3.2-fold at 60-80 degrees C, reflecting the thermal stability of the enzyme. The reaction required no metal ion and was activated slightly by low concentrations (1-5 mM) of EDTA. The apparent K(m) value for sepiapterin was determined as 0.92 mM and the V(max) value was 151.3 nmol/min/mg. The new catalytic function of E. coli PTPS does not imply any physiological role, because sepiapterin is not an endogenous substrate of the organism. The same activity, however, was also detected in a PTPS ortholog of Synechocystis sp. PCC 6803 but not significant in Drosophila and human enzymes, suggesting that the activity may be prevalent in bacterial PTPS orthologs.

Production of sepiapterin in Escherichia coli by coexpression of cyanobacterial GTP cyclohydrolase I and human 6-pyruvoyltetrahydropterin synthase.

Synechocystis sp. strain PCC 6803 GTP cyclohydrolase I and human 6-pyruvoyltetrahydropterin synthase were coexpressed in Escherichia coli. The E. coli transformant produced sepiapterin, which was identified by high-performance liquid chromatography and enzymatically converted to dihydrobiopterin by sepiapterin reductase. Aldose reductase, another indispensable enzyme for sepiapterin production, may be endogenous in E. coli.