TY - JOUR
T1 - Direct Enzymatic Synthesis of 1-Phosphatidyl-β-D-glucose by Engineered Phospholipase D
AU - Inoue, Arisa
AU - Adachi, Masaatsu
AU - Damnjanović, Jasmina
AU - Nakano, Hideo
AU - Iwasaki, Yugo
N1 - Funding Information:
This work was supported by a Grant-in-Aid for scientific research C from the Japan Society for the Promotion of Science to Y.I. (25420830).
PY - 2016/8/16
Y1 - 2016/8/16
N2 - A simple, direct method for the synthesis of 1-phosphatidyl-β-D-glucose (1-PGlc) was established, being the first example of direct enzymatic synthesis of this recently discovered, highly important bioactive phospholipid. The method employs phospholipase D (PLD, E.C. 3.1.4.4)-catalyzed transphosphatidylation, in which the polar head group of phosphatidylcholine is exchanged to glucose. Although wild-type PLD can catalyze the transfer reaction, it provides only 6-phosphatidyl-glucose, a positional isomer of 1-PGlc, due to its strong preference towards the primary hydroxyl group. To synthesize 1-PGlc, engineered PLD variants, previously isolated as the ones active on secondary hydroxyls of inositol, were screened for the ability to catalyze the reaction. One of the variants, 187K/191W/385Y (KWY), was identified as the best-performing in 1-PGlc synthesis, however, in addition to 1-PGlc the reaction with KWY also afforded undesired positional isomers as byproducts. To facilitate the isolation of 1-PGlc, the isomers were converted into the corresponding amines by reductive amination. Following the column chromatography, the desired product was isolated with the overall yield of 12.5 %. The structure of the product was confirmed by 1H-NMR analysis.
AB - A simple, direct method for the synthesis of 1-phosphatidyl-β-D-glucose (1-PGlc) was established, being the first example of direct enzymatic synthesis of this recently discovered, highly important bioactive phospholipid. The method employs phospholipase D (PLD, E.C. 3.1.4.4)-catalyzed transphosphatidylation, in which the polar head group of phosphatidylcholine is exchanged to glucose. Although wild-type PLD can catalyze the transfer reaction, it provides only 6-phosphatidyl-glucose, a positional isomer of 1-PGlc, due to its strong preference towards the primary hydroxyl group. To synthesize 1-PGlc, engineered PLD variants, previously isolated as the ones active on secondary hydroxyls of inositol, were screened for the ability to catalyze the reaction. One of the variants, 187K/191W/385Y (KWY), was identified as the best-performing in 1-PGlc synthesis, however, in addition to 1-PGlc the reaction with KWY also afforded undesired positional isomers as byproducts. To facilitate the isolation of 1-PGlc, the isomers were converted into the corresponding amines by reductive amination. Following the column chromatography, the desired product was isolated with the overall yield of 12.5 %. The structure of the product was confirmed by 1H-NMR analysis.
KW - enzyme catalysis
KW - phospholipase D, phosphatidylglucoside
KW - phospholipid
KW - transphosphatidylation
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U2 - 10.1002/slct.201600839
DO - 10.1002/slct.201600839
M3 - Article
AN - SCOPUS:85041961331
VL - 1
SP - 4121
EP - 4125
JO - ChemistrySelect
JF - ChemistrySelect
SN - 2365-6549
IS - 13
ER -