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.
Seo
Jiyoung
J
Department of Environmental Science and Engineering, Gwangju Institute of Science and Technology, Gwangju 500-712, Republic of Korea.
Kang
Su-Il
SI
Kim
Mihyang
M
Won
Dongho
D
Takahashi
Haruko
H
Ahn
Joong-Hoon
JH
Chong
Youhoon
Y
Lee
Eunjung
E
Lim
Yoongho
Y
Kanaly
Robert A
RA
Han
Jaehong
J
Hur
Hor-Gil
HG
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