Conversion of pipecolic acid into lysine in Penicillium chrysogenum requires pipecolate oxidase and saccharopine reductase: Characterization of the lys7 gene encoding saccharopine reductase

Pipecolic acid is a component of several secondary metabolites in plants an d fungi. This compound is useful as a precursor of nonribosomal peptides wi th novel pharmacological activities. In Penicillium chrysogenum pipecolic a cid is converted into lysine and complements the lysine requirement of thre e different lysine auxotrophs with mutations in the lys1, lys2, or lys3 gen es allowing a slow growth of these auxotrophs. We have isolated two P. chry sogenum mutants, named 7.2 and 10.25, that are unable to convert pipecolic acid into lysine. These mutants lacked, respectively, the pipecolate oxidas e that converts pipecolic acid into piperideine-6-carboxylic acid and the s accharopine reductase that catalyzes the transformation of piperideine-6-ca rboxylic acid into saccharopine. The 10.25 mutant was unable to grow in Cza pek medium supplemented with a-aminoadipic acid. A DNA fragment complementi ng the 10.25 mutation has been cloned; sequence analysis of the cloned gene (named lys7) revealed that it encoded a protein with high similarity to th e saccharopine reductase from Neurospora crassa, Magnaporthe grisea, Saccha romyces cerevisiae, and Schizosaccharomyces pombe. Complementation of the 1 0.25 mutant with the cloned gene restored saccharopine reductase activity, confirming that lys7 encodes a functional saccharopine reductase. Our data suggest that in P. chrysogenum the conversion of pipecolic acid into lysine proceeds through the transformation of pipecolic acid into pip erideine-6- carboxylic acid, saccharopine, and lysine by the consecutive action of pipe colate oxidase, saccharopine reductase, and saccharopine dehydrogenase.