Identification of covalent binding sites of phthalic anhydride in human hemoglobin.
Phthalic anhydride (PA) is a reactive low molecular weight compound used in the chemical industry. The exposure of PA may lead to work-related airway diseases such as rhinitis, chronic bronchitis, and asthma. The exposure gives rise to an increase in hapten-specific IgG antibodies in workers but with a low presence of specific IgE antibodies. In this study, the binding of PA to human hemoglobin (Hb) in vitro was investigated. Trypsin and Pronase E digestion, LC, LC/MS/MS, GC/MS analysis, and nanoelectrospray hybrid quadrupole time-of-flight MS were used to identify the adducted amino acids of the synthesized PA-Hb conjugates. In the conjugate with the molar ratio 1:0.1, a total of six adducted amino acids were identified. N-Terminal valine was found adducted in both the alpha- and the beta-chains as well as a total of four lysines, Val 1, Lys 16, and Lys 61 on the alpha-chain and Val 1, Lys 66, and Lys 144 on the beta-chain. Two types of lysine adducts were found, a phthalamide and a phthalimide. It was also found that PA differs in its binding site as compared to hexahydrophthalic anhydride. The result of this study suggests several interesting applications of biological monitoring.
Jeppsson
Marina C
MC
Department of Occupational and Environmental Medicine, Institute of Laboratory Medicine, University Hospital, SE-221 85 Lund, Sweden. Marina.Jeppsson@med.lu.se
Jönsson
Bo A G
BA
Kristiansson
Monica
M
Lindh
Christian H
CH
D000595:Amino Acid Sequence; D001665:Binding Sites; D002853:Chromatography, Liquid; D006454:Hemoglobins / Q000737:chemistry* / Q000378:metabolism; D006801:Humans; D006868:Hydrolysis; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D010796:Phthalic Anhydrides / Q000737:chemistry* / Q000378:metabolism; D011402:Pronase / Q000378:metabolism; D011485:Protein Binding; D014357:Trypsin / Q000378:metabolism
D006454:Hemoglobins; D010796:Phthalic Anhydrides; D014357:Trypsin; D011402:Pronase