Data di Pubblicazione:
2011
Citazione:
Characterization of albumin carbonylation by a tandem ms-precursor ion approach / M. Orioli, A. Pancotti, M. Scognamiglio, M. Carini, G. Aldini - In: Atti del congresso: 14th Recent Development in Pharmaceutical Analysis 2011[s.l] : Divisione di Chimica Farmaceutica, 2011. (( Intervento presentato al 14. convegno Recent Development in Pharmaceutical Analysis tenutosi a Pavia nel 2011.
Abstract:
Several experimental evidences confirm that protein carbonylation induced by
reactive carbonyl species (RCS) is involved in the pathogenesis of several oxidative
based diseases, including atherosclerosis and diabetic related diseases. By using a
proteomic approach, we previously found that human serum albumin (HSA) is the
main nucleophilic target of human plasma, due to the presence of some reactive
nucleophilic aminoacid residues. We here report the set-up of a liquid
chromatography electrospray ionisation multi-stage mass spectrometry (LC-ESIMS/
MS) approach based on precursor-ion scanning, aimed to identify and
characterize the carbonylated adducts on His146, a reactive nucleophilic site of HSA
able to form Michael adducts with alfa,beta-unsaturated aldehydes. The first step
consisted of identifying the product ions to be used in the precursor ion scan, relative
to the His146 peptide generated by the enzymatic digestion. In particular, by
considering the trypsin/chymotrypsin digested peptide containing His146
(HPYFYAPELLFFAK), the ions at m/z 625.2 (y5) and m/z 964.6 (y8) were selected
as product ions since not containing the target nucleophilic residue. MS data were
then analyzed by using an algorithm developed by us, permitting a rapid and specific
identification of the precursor-ions attributed to native and modified peptides. In a
following step, the identified precursor ions were selected for product ion scan
analysis in order to identify and characterize the covalent modification. The method
has been validated by using 4-hydroxy-trans-nonenal (HNE) as a RCS model.
Human serum was incubated in the presence of HNE (10 nmoles/ml), and then HSA
was isolated by affinity chromatography and digested by using trypsin/chymotrypsin.
The approach identified two peaks at m/z 871.9 [M+2H]2+ and m/z 949.9 [M+2H]2+
relative to the HPYFYAPELLFFAK, the former attributed to the native peptide and the
latter to the His146-HNE adduct, as confirmed by MS/MS. The method was then
successfully applied for the identification and characterization of the His146 covalent
adducts of HSA incubated with 4-hydroxy-hexenal, crotonaldehyde, nonenale and
acrolein. The tandem MS approach here reported could be a powerful tool for the
rapid identification and characterization of carbonylated albumin in different
pathological conditions.
Tipologia IRIS:
03 - Contributo in volume
Elenco autori:
M. Orioli, A. Pancotti, M. Scognamiglio, M. Carini, G. Aldini
Link alla scheda completa:
Titolo del libro:
Atti del congresso: 14th Recent Development in Pharmaceutical Analysis 2011