Advanced glycation end products of beta2-microglobulin in uremic patients as determined by high resolution mass spectrometry.
By using a high resolution top-down and bottom-up approach we identified and characterized the AGEs of beta2-microglobulin (β2-m) formed by incubating the protein in the presence of glucose and of the main reactive carbonyl species. Glucose induced glycation on the N-terminal residue, while glyoxal (GO) and methylglyoxal (MGO) covalently reacted with Arg3. Carboxymethyl (CM-R) and imidazolinone (R-GO) derivatives were identified in the case of GO and carboxyethyl arginine (CE-R) and methyl-imidazolinone (R-MGO) for MGO. Interestingly, α,β-unsaturated aldehydes [4-hydroxy-2-nonenal (HNE); 4-oxo-2-nonenal (ONE); acrolein (ACR)] did not induce any covalent modifications up to 100μM. The different reactivity of β2-m towards the different RCS was then rationalized by molecular modeling studies. The MS method was then applied to fully characterize the AGEs of β2-m isolated from the urine of uremic subjects. CM-R, CE-R and R-MGO were easily identified on Arg3 and their relative abundance in respect to the native protein determined by a semi-quantitative approach. Overall, the AGEs content of urinary β2-m ranged from 0.2 to 1% in uremic subjects. The results here reported offer novel insights and technical achievements for a potential biological role of AGEs-β2-m in pathological conditions.
Department of Drug Sciences, University of Pavia, Viale Taramelli 12, 27100 Pavia, Italy. Electronic address: laura.bertoletti@unipv.it.
Laura
L
Bertoletti
Department of Pharmaceutical Sciences, Università degli Studi di Milano, Via Mangiagalli 25, 20133 Milan, Italy. Electronic address: lregazzoni@berkeley.edu.
Luca
L
Regazzoni
Department of Pharmaceutical Sciences, Università degli Studi di Milano, Via Mangiagalli 25, 20133 Milan, Italy. Electronic address: altomare.alessandra@unimi.it.
Alessandra
A
Altomare
Department of Drug Sciences, University of Pavia, Viale Taramelli 12, 27100 Pavia, Italy. Electronic address: raffaella.colombo@unipv.it.
Raffaella
R
Colombo
Department of Pharmaceutical Sciences, Università degli Studi di Milano, Via Mangiagalli 25, 20133 Milan, Italy. Electronic address: mara.colzani@unimi.it.
Mara
M
Colzani
Department of Pharmaceutical Sciences, Università degli Studi di Milano, Via Mangiagalli 25, 20133 Milan, Italy. Electronic address: giulio.vistoli@unimi.it.
Giulio
G
Vistoli
Department of Molecular Medicine, University of Pavia, Via Forlanini 6, 27100 Pavia, Italy. Electronic address: loredana.marchese@unipv.it.
Loredana
L
Marchese
Department of Pharmaceutical Sciences, Università degli Studi di Milano, Via Mangiagalli 25, 20133 Milan, Italy. Electronic address: marina.carini@unimi.it.
Marina
M
Carini
Department of Drug Sciences, University of Pavia, Viale Taramelli 12, 27100 Pavia, Italy. Electronic address: ersidelo@unipv.it.
Ersilia
E
De Lorenzi
Department of Pharmaceutical Sciences, Università degli Studi di Milano, Via Mangiagalli 25, 20133 Milan, Italy. Electronic address: giancarlo.aldini@unimi.it.
Giancarlo
G
Aldini
D000171:Acrolein / Q000737:chemistry; D000447:Aldehydes / Q000737:chemistry; D001120:Arginine / Q000737:chemistry; D005947:Glucose / Q000737:chemistry; D017127:Glycation End Products, Advanced / Q000737:chemistry* / Q000652:urine*; D006037:Glyoxal / Q000737:chemistry; D006801:Humans; D013058:Mass Spectrometry / Q000379:methods; D011765:Pyruvaldehyde / Q000737:chemistry; D014511:Uremia / Q000378:metabolism / Q000652:urine*; D001613:beta 2-Microglobulin / Q000378:metabolism*
C403894:4-oxo-2-nonenal; D000447:Aldehydes; D017127:Glycation End Products, Advanced; D001613:beta 2-Microglobulin; D006037:Glyoxal; D011765:Pyruvaldehyde; D000171:Acrolein; D001120:Arginine; D005947:Glucose; C027576:4-hydroxy-2-nonenal
Advanced glycoxidation end products; Beta(2)-microglobulin; Dialysis related amyloidosis; Mass spectrometry; Oxidative stress; Reactive carbonyl species