Reactions involved in the conversion of ornithine to proline in Clostridia.
An enzyme system which converts ornithine to proline was partially purified from extracts of cells of Clostridium botulinum and of Clostridium PA 3670 by fractionation with ammonium sulfate and by dialysis in the presence of 0.01 m ornithine. Nicotinamide adenine dinucleotide (NAD) was the only cofactor required for maximal activity of the partially purified system. A possible intermediate in the conversion was accumulated when a high concentration of proline was used as substrate and the NAD was maintained in the oxidized state by adding lactic dehydrogenase. Small but significant amounts of this or a similar compound were trapped with either ornithine or proline as substrate when o-aminobenzaldehyde was added to reaction mixtures. The accumulation of the o-aminobenzaldehyde reaction product was NAD-dependent with both substrates. The compound accumulated from proline was identified as Delta(1)-pyrroline-5-carboxylic acid on the basis of the melting point of the 2,4-dinitrophenylhydrazone, and by paper chromatography of the reaction product formed with o-aminobenzaldehyde. Also, extracts of C. botulinum cells oxidized reduced NAD (NADH) in the presence of the product from proline or in the presence of Delta(1)-pyrroline-5-carboxylic acid, but did not do so in the presence of the other possible intermediate, Delta(1)-pyrroline-2-carboxylic acid. (14)C-Delta(1)-pyrroline-5-carboxylic acid was reduced to (14)C-proline by these extracts in the presence of NADH. These data indicate that the conversion of ornithine to proline by C. botulinum and Clostridium PA 3679 cells involves an oxidation of ornithine to glutamic-gamma-semialdehyde which undergoes ring closure to Delta(1)-pyrroline-5-carboxylic acid. The latter compound is then reduced to proline.
D002247:Carbon Isotopes; D003013:Clostridium / Q000201:enzymology / Q000378:metabolism*; D003014:Clostridium botulinum / Q000378:metabolism; D009243:NAD / Q000378:metabolism; D009952:Ornithine / Q000378:metabolism*; D010084:Oxidation-Reduction; D011392:Proline / Q000096:biosynthesis*; D013057:Spectrum Analysis
D002247:Carbon Isotopes; D009243:NAD; D011392:Proline; D009952:Ornithine