Accumulation of the secondary metabolite precursors piperideine-6-carboxylic acid and pipecolic acid by disruption of lys7 gene encoding saccharopine reductase in Penicillium chrysogenum
Naranjo,
L.; Martín de Valmaseda, E.; Casqueiro, J.; V. Ullán,
R.; Lamas, M.; Bañuelos, O. y Martín, J.F.
Área
de Microbiología, Facultad de Biología, Universidad de León,
24071 León,
Instituto de biotecnología (INBIOTEC), Avda. del Real Nº1,
24006 León.
Pipecolic acid serves as precursor of the biosynthesis of the alkaloids slaframine and swainsonine (an antitumor agent) in some fungi. It is not known whether other fungi are able to synthesize pipecolic acid. Penicillium chrysogenum has a very active a-aminoadipic acid pathway that is used for synthesis of this precursor of penicillin. The lys7 gene encoding saccharopine reductase of P. chrysogenum was target-inactivated by the double recombination method. Analysis of disrupted strain (named P. chrysogenum SR1-) showed the presence of a mutant lys7 gene lacking about 1.000 bp in the 3´-end region. The P. chrysogenum SR1- strain lacked saccharopine reductase activity, which was recovered after transformation of this mutant with the intact lys7 gene in an autonomous replicating plasmid. The P. chrysogenum SR1- strain was a lysine auxotroph and accumulated P6C. When the P. chrysogenum SR1- mutant was grown with L-lysine as sole nitrogen source and supplemented with D,L-a-aminoadipic acid, a high level of pipecolic acid accumulated intracellularly. This mutant strain is therefore, useful for the production of pipecolate-containing secondary metabolites. A comparison of the SR1- strain with a lys2 defective mutant provided evidence showing that P. chrysogenum synthesizes pipecolic acid from a-aminoadipic acid and not from lysine catabolism.
