Biotransformation of L-lysine to L-pipecolic acid catalyzed by L-lysine 6-aminotransferase and pyrroline-5-carboxylate reductase.
The enzyme involved in the reduction of delta1-piperideine-6-carboxylate (P6C) to L-pipecolic acid (L-PA) has never been identified. We found that Escherichia coli JM109 transformed with the lat gene encoding L-lysine 6-aminotransferase (LAT) converted L-lysine (L-Lys) to L-PA. This suggested that there is a gene encoding "P6C reductase" that catalyzes the reduction of P6C to L-PA in the genome of E. coli. The complementation experiment of proC32 in E. coli RK4904 for L-PA production clearly shows that the expression of both lat and proC is essential for the biotransformation of L-Lys to L-PA. Further, We showed that both LAT and pyrroline-5-carboxylate (P5C) reductase, the product of proC, were needed to convert L-Lys to L-PA in vitro. These results demonstrate that P5C reductase catalyzes the reduction of P6C to L-PA. Biotransformation of L-Lys to L-PA using lat-expressing E. coli BL21 was done and L-PA was accumulated in the medium to reach at an amount of 3.9 g/l after 159 h of cultivation. It is noteworthy that the ee-value of the produced pipecolic acid was 100%.
Fujii
Tadashi
T
Bioresource Laboratories, Mercian Corp, Johnan, Fujisawa, Japan. fujii-td@mercian.co.jp
Mukaihara
Manabu
M
Agematu
Hitosi
H
Tsunekawa
Hiroshi
H
D002851:Chromatography, High Pressure Liquid; D002853:Chromatography, Liquid; D002855:Chromatography, Thin Layer; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism*; D005285:Fermentation; D005816:Genetic Complementation Test; D051306:L-Lysine 6-Transaminase; D008239:Lysine / Q000378:metabolism*; D013058:Mass Spectrometry; D010084:Oxidation-Reduction; D010875:Pipecolic Acids / Q000378:metabolism*; D010957:Plasmids / Q000235:genetics; D011762:Pyrroline Carboxylate Reductases / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry; D000637:Transaminases / Q000235:genetics / Q000378:metabolism*
D010875:Pipecolic Acids; D011994:Recombinant Proteins; D011762:Pyrroline Carboxylate Reductases; D000637:Transaminases; D051306:L-Lysine 6-Transaminase; C031345:pipecolic acid; D008239:Lysine