Efficient synthesis of trans-polyisoprene compounds using two thermostable enzymes in an organic-aqueous dual-liquid phase system.
The trans-polyisoprene compounds are synthesized by trans-isoprenyl diphosphate synthase (IDS) with consecutive condensation of isopentenyl diphosphate (IPP) to dimethylallyl diphosphate (DMAPP). The in vitro condensation by IDS does not proceed efficiently by hydrophobic interaction between IDS and the hydrocarbon of longer products. In the present study, the enzymatic synthesis of trans-polyisoprenyl diphosphates was attempted in an organic-aqueous dual-liquid phase system with thermostable enzymes obtained from Thermococcus kodakaraensis. The conversion from DMAPP to a longer-chain product was achieved in a dual-liquid phase system, and more than 80% of the products were recovered in the organic phase. When the mutant IDS-Y81S, in which Tyr81 is replaced with Ser, was used in the dual-phase system, productivity was enhanced about four times and the ratio of the longer-chain products was increased. Co-incubation of IPP isomerase from T. kodakaraensis with IDS or IDS-Y81S enabled the direct synthesis of polyisoprenyl diphosphates from IPPs.
Fujiwara
Shinsuke
S
Department of Bioscience, Nanobiotechnology Research Center, School of Science and Technology, Kwansei Gakuin University, 2-1 Gakuen, Sanda 699-1337, Japan. fujiwara-s@kwansei.ac.jp
Yamanaka
Aiko
A
Yamada
Yosuke
Y
Hirooka
Kazutake
K
Higashibata
Hiroki
H
Fukuda
Wakao
W
Nakayama
Jun-ichi
J
Imanaka
Tadayuki
T
Fukusaki
Ei-ichiro
E
D019843:Archaeal Proteins / Q000378:metabolism; D019748:Carbon-Carbon Double Bond Isomerases / Q000378:metabolism; D002942:Circular Dichroism; D045782:Hemiterpenes / Q000378:metabolism; D007536:Isomerism; D008954:Models, Biological; D009943:Organophosphorus Compounds / Q000378:metabolism; D044367:Phase Transition; D011106:Polyisoprenyl Phosphates / Q000096:biosynthesis*; D013696:Temperature; D019712:Thermococcus / Q000201:enzymology*
D019843:Archaeal Proteins; D045782:Hemiterpenes; D009943:Organophosphorus Compounds; D011106:Polyisoprenyl Phosphates; C004809:isopentenyl pyrophosphate; C043060:3,3-dimethylallyl pyrophosphate; D019748:Carbon-Carbon Double Bond Isomerases; C021453:isopentenyldiphosphate delta-isomerase