Reductive catabolism of pyrimidine bases by Pseudomonas stutzeri

Abstract

Pyrimidine base catabolism in Pseudomonas stutzeri ATCC 17588 was investigated and was found to occur by means of the reductive pathway. Pyrimidine bases and their respective reductive pathway catabolic products could serve as nitrogen sources for growth of P. stutzeri. Activities of the three enzymes associated with the reductive pathway of pyrimidine catabolism were detected in cells of P. stutzeri. The initial enzyme of the reductive pathway, dihydropyrimidine dehydrogenase, utilized NADH as its nicotinamide cofactor. Cells grown on pyrimidine bases as nitrogen sources contained elevated dehydrogenase activity relative to those grown on ammonium sulphate as nitrogen source. Activities of the second and third reductive pathway enzymes, dihydropyrimidinase and N-carbamoyl-P-alanine amidohydrolase, respectively, were also affected by growth conditions. If pyrimidine or dihydropyrimidine bases served as nitrogen sources, increases in the levels of these enzymes were observed compared to their activities determined when the nitrogen source was ammonium sulphate.

Reductive catabolism of uracil and thymine by Burkholderia cepacia

“…In cells grown on uracil or thymine as a nitrogen source, no dehydrogenase activity [< 0.02 ± 0.01 or < 0.04 ± 0.01 nmol product min -1 (mg protein) -1 , respectively] was detected using 0.2 mM NADH and either pyrimidine base (1 mM) as an enzyme substrate. This finding differs from previous studies exploring pyrimidine catabolism in the closely related pseudomonads where the preferred nicotinamide cofactor for the dehydrogenase was NADH (Kim and West 1991;West 1991;Xu and West 1992). In B. pickettii ATCC 27511, dehydrogenase activity was fivefold higher using NADH instead of NADPH as the cofactor in extracts from succinate-grown cells (West 1994).…”

“…The highest amidohydrolase activity was seen in glucose-grown cells that used thymine as a nitrogen source, while β-alanine or β-aminoisobutyric acid affected amidohydrolase activity to a lesser extent (Table 1). In the closely related pseudomonads, the reductive catabolic enzyme activities have also been shown to be affected by growth on pyrimidine and dihydropyrimidine bases as nitrogen sources (Kim and West 1991;West 1991;Xu and West 1992).…”

“…The activity of dihydropyrimidinase was determined using a colorimetric assay for Ncarbamoyl-β-alanine that has been previously described (Xu and West 1992). N-carbamoyl-β-alanine amidohydrolase was assayed by following the release of ammonium ions during the reaction (Xu and West 1992). Protein was measured by the method of Bradford (1976), and lysozyme served as the standard protein.…”

“…Dihydropyrimidine dehydrogenase was assayed with NADPH at 340 nm using a previously described protocol (West 1991). The activity of dihydropyrimidinase was determined using a colorimetric assay for Ncarbamoyl-β-alanine that has been previously described (Xu and West 1992). N-carbamoyl-β-alanine amidohydrolase was assayed by following the release of ammonium ions during the reaction (Xu and West 1992).…”

“…The reductive pathway culminates with β-alanine or β-aminoisobutyric acid being produced from their respective N-carbamoyl derivatives by the enzyme N-carbamoyl-β-alanine amidohydrolase (EC 3.5.1.6). The reductive pathway is known to be active in a number of prokaryotic species (Vogels and van der Drift 1976;Kim and West 1991;West 1991;Xu and West 1992;West 1994).…”