Reaction of aflatoxin B(1) oxidation products with lysine.
Aflatoxin (AF) B(1) exo-8,9-epoxide hydrolysis yields AFB(1) dihydrodiol, which undergoes base-catalyzed rearrangement to, and is in equilibrium with, AFB(1) dialdehyde. We investigated the reaction of AFB(1) dialdehyde with albumin to generate a Lys adduct, previously characterized by others [Sabbioni, G., Skipper, P. L., Büchi, G., and Tannenbaum, S. R. (1987) Carcinogenesis8, 819-824; Sabbioni, G. (1990) Chem.-Biol. Interact. 75, 1-15]. Pronase digestion of bovine albumin serum treated with AFB(1) dialdehyde and HPLC yielded the adduct, identified by its characteristic UV and mass spectra. The structure of the Lys-AFB(1) dialdehyde adduct is concluded to be (S)-alpha-amino-2,3-dihydro-2-oxo-4-(1,2,3,4-tetrahydro-7-hydroxy-9-methoxy-3,4-dioxocyclopenta[c][1]benzopyran-6-yl)-1H-pyrrole-1-hexanoic acid, structure B of the former paper and 8 of the latter, based on work with the methylamine adduct described in the following paper in this issue [Guengerich, F. P., Voehler, M., Williams, K. M., Deng, Z., and Harris, T. M. (2002) Chem. Res. Toxicol. 15, 793-798]. The time course of product formation at varying concentrations of AFB(1) dialdehyde could be described by complexation with albumin with a K(d) of 1.5 mM and a first-order reaction rate with the N6-amino group of Lys of 0.033 min(-)(1). The reaction of AFB(1) dialdehyde with N(2)-acetylLys was monitored by UV spectroscopy and yielded a final spectrum similar to that of the described Lys adduct. Kinetic analysis of the changes at pH 7.2 was best described with a scheme involving equilibrium of the dialdehyde with dihydrodiol and a rate-limiting reaction of AFB(1) dialdehyde with the N6 atom of N(2)-acetylLys, with an apparent second-order rate constant of 2.6 x 10(3) M(-)(1) min(-)(1), followed by putative carbinolamine formation and rearrangement, collectively described by a first-order rate constant of 7.6 min(-)(1). Competition experiments with the hydrolysis of AFB(1) exo-8,9-epoxide indicate that N2-acetylLys also reacts with the epoxide at pH 7.2 (k = 350 M(-)(1) min(-)(1)) and 9.5 (k = 1.8 x 10(3) M(-)(1) min(-)(1)). This reaction might contribute to the formation of protein Lys adducts, depending upon the local concentration of free or protein Lys. Mass spectral analysis of trypsin digests of bovine serum albumin modified with AFB(1) dialdehyde indicated selective modification of Lys455 and Lys548. Collectively, these results provide more insight into the mechanism of formation of AFB(1) dialdehyde-protein adducts and indicate that the formation of Lys adducts is a moderately efficient process. The binding of AFB(1) dialdehyde to albumin or the protonation of the N6-amino group retards the reaction with Lys residues.
Guengerich
F Peter
FP
Department of Biochemistry and Center in Molecular Toxicology, Vanderbilt University, Nashville, Tennessee 37232-0146, USA. guengerich@toxicology.mc.vanderbilt.edu
Arneson
Kyle O
KO
Williams
Kevin M
KM
Deng
Zhengwu
Z
Harris
Thomas M
TM
D016604:Aflatoxin B1 / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism*; D000447:Aldehydes / Q000737:chemistry / Q000378:metabolism; D000818:Animals; D001665:Binding Sites; D001667:Binding, Competitive; D002021:Buffers; D002417:Cattle; D002851:Chromatography, High Pressure Liquid; D005978:Glutathione / Q000031:analogs & derivatives / Q000737:chemistry; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008239:Lysine / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism*; D010084:Oxidation-Reduction; D012709:Serum Albumin / Q000378:metabolism; D012710:Serum Albumin, Bovine / Q000737:chemistry / Q000378:metabolism*; D021241:Spectrometry, Mass, Electrospray Ionization; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013056:Spectrophotometry, Ultraviolet
D000447:Aldehydes; D002021:Buffers; D012709:Serum Albumin; C052669:aflatoxin B1-bovine serum albumin; C002315:N(alpha)-acetyllysine; D012710:Serum Albumin, Bovine; C006426:aflatoxin B1-2,3-oxide; D016604:Aflatoxin B1; D005978:Glutathione; D008239:Lysine