Time-dependent density functional theory-assisted absolute configuration determination of cis-dihydrodiol metabolite produced from isoflavone by biphenyl dioxygenase.
Escherichia coli cells containing the biphenyl dioxygenase genes bphA1A2A3A4 from Pseudomonas pseudoalcaligenes KF707 were found to biotransform isoflavone and produced a metabolite that was not found in a control experiment. Liquid chromatography/mass spectrometry (LC/MS) and (1)H and (13)C nuclear magnetic resonance (NMR) analyses indicated that biphenyl dioxygenase induced 2',3'-cis-dihydroxylation of the B-ring of isoflavone. In a previous report, the same enzyme showed dioxygenase activity toward flavone, producing flavone 2',3'-cis-dihydrodiol. Due to growing interest in flavone chemistry and the absolute configuration of natural products, time-dependent density functional theory (TD-DFT) calculations were combined with circular dichroism (CD) spectroscopy to determine the absolute configuration of the isoflavone dihydrodiol. By computational methods, the structure of the isoflavone metabolite was determined to be 3-[(5S,6R)-5,6-dihydroxycyclohexa-1,3-dienyl]-4H-chromen-4-one. This structure was confirmed further by the modified Mosher's method. The same protocol was applied to the flavone metabolite, and the absolute configuration was determined to be 2-[(5S,6R)-5,6-dihydroxycyclohexa-1,3-dienyl]-4H-chromen-4-one. After determination of the absolute configurations of the biotransformation products, we suggest the binding mode of these substrate analogs to the enzyme active site.
Department of Environmental Science and Engineering, Gwangju Institute of Science and Technology, Gwangju 500-712, Republic of Korea.
Jiyoung
J
Seo
Su-Il
SI
Kang
Mihyang
M
Kim
Dongho
D
Won
Haruko
H
Takahashi
Joong-Hoon
JH
Ahn
Youhoon
Y
Chong
Eunjung
E
Lee
Yoongho
Y
Lim
Robert A
RA
Kanaly
Jaehong
J
Han
Hor-Gil
HG
Hur
D001665:Binding Sites; D001711:Biotransformation; D020134:Catalytic Domain; D002942:Circular Dichroism / Q000379:methods*; D003198:Computer Simulation; D003511:Cyclohexanols / Q000737:chemistry; D049308:Dioxygenases / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D007529:Isoflavones / Q000737:chemistry / Q000378:metabolism*; D007536:Isomerism; D009682:Magnetic Resonance Spectroscopy / Q000379:methods*; D008968:Molecular Conformation; D009281:Naphthalenes / Q000737:chemistry*; D044205:Pseudomonas pseudoalcaligenes / Q000201:enzymology / Q000235:genetics; D013997:Time Factors
D003511:Cyclohexanols; D007529:Isoflavones; D009281:Naphthalenes; C507866:trans-1,2-dihydro-1,2-naphthalenediol; C046203:5,6-dihydroxycyclohexa-1,3-diene; D049308:Dioxygenases