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Identification and relative quantification of specific glycation sites in human serum albumin

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Abstract

Glycation (or non-enzymatic glycosylation) is a common non-enzymatic covalent modification of human proteins. Glucose, the highest concentrated monosaccharide in blood, can reversibly react with amino groups of proteins to form Schiff bases that can rearrange to form relatively stable Amadori products. These can be further oxidized to advanced glycation end products (AGEs). Here, we analyzed the glycation patterns of human serum albumin (HSA) in plasma samples obtained from five patients with type 2 diabetes mellitus. Therefore, glycated peptides from a tryptic digest of plasma were enriched with m-aminophenylboronic acid (mAPBA) affinity chromatography. The glycated peptides were then further separated in the second dimension by RP-HPLC coupled on-line to an electrospray ionization (ESI) tandem mass spectrometer (MS/MS). Altogether, 18 Amadori peptides, encompassing 40% of the HSA sequence, were identified. The majority of the peptides were detected and relatively quantified in all five samples with a high reproducibility among the replicas. Eleven Lys-residues were glycated at similar quantities in all samples, with glycation site Lys549 (KAm(Glc)QTALVELVK) being the most abundant. In conclusion, the established mAPBA/nanoRP-HPLC-ESI-MS/MS approach could reproducibly identify and quantify glycation sites in plasma samples, potentially useful in diagnosis and therapeutic control.

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Abbreviations

AGE:

advanced glycation end product

CID:

collision-induced dissociation

CEL:

Nε-carboxyethyllysine

CML:

Nε-carboxymethyllysine

2D-LC:

two-dimensional liquid chromatography

DTT:

dithiothreitol

ESI:

electrospray ionization

glycBSA:

glycated bovine serum albumin

HbA1c :

glycated hemoglobin fraction

HSA:

human serum albumin

IDA:

information-dependent acquisition

MALDI:

matrix-assisted laser desorption/ionization

mAPBA:

m-aminophenylboronic acid

MS:

mass spectrometry

MS/MS:

tandem mass spectrometry

SDS-PAGE:

polyacrylamide gel electrophoresis with sodium dodecyl sulfate

QqTOF:

quadrupole time-of-flight

RPC:

reversed-phase chromatography

RP-HPLC:

reversed-phase high-performance liquid chromatography

TIC:

total ion current

XIC:

extracted ion chromatogram

References

  1. Gustin MC, Albertyn J, Alexander M, Davenport K (1998) Microbiol Mol Biol Rev 62:1264–1300

    CAS  Google Scholar 

  2. Dennis JW, Granovsky M, Warren CE (1999) Bioessays 21:412–421

    Article  CAS  Google Scholar 

  3. Risteli J, Kivirikko KI (1974) Biochem J 144:115–122

    CAS  Google Scholar 

  4. Wade PA, Pruss D, Wolffe AP (1997) Trends Biochem Sci 22:128–132

    Article  CAS  Google Scholar 

  5. Cloos PA, Christgau S (2002) Matrix Biol 21:39–52

    Article  CAS  Google Scholar 

  6. Ulrich P, Cerami A (2001) Recent Prog Horm Res 56:1–21

    Article  CAS  Google Scholar 

  7. Maillard LC, Gautier MA (1912) Compte-rendu de l'Académie des sciences 154:66–68

    CAS  Google Scholar 

  8. Pilkova L, Pokorny J, Davidek J (1990) Die Nahrung 34:759–764

    Article  CAS  Google Scholar 

  9. Grillo MA, Colombatto S (2007) Amino Acids 35:29–36

    Article  CAS  Google Scholar 

  10. Tierney LM, McPhee SJ, Papadakis MA (2001) Current medical diagnosis and treatment 2002. McGraw-Hill, London

    Google Scholar 

  11. Alberti KG, Zimmet PZ (1998) Diabet Med 15:539–553

    Article  CAS  Google Scholar 

  12. Dyer DG, Dunn JA, Thorpe SR, Bailie KE, Lyons TJ, McCance DR, Baynes JW (1993) J Clin Invest 91:2463–2469

    Article  CAS  Google Scholar 

  13. Miyazawa N, Kawasaki Y, Fujii J, Theingi M, Hoshi A, Hamaoka R, Matsumoto A, Uozumi N, Teshima T, Taniguchi N (1998) Biochem J 336(Pt 1):101–107

    CAS  Google Scholar 

  14. Ahmed N, Babaei-Jadidi R, Howell SK, Thornalley PJ, Beisswenger PJ (2005) Diabetes Care 28:2465–2471

    Article  CAS  Google Scholar 

  15. Gabbay KH, Hasty K, Breslow JL, Ellison RC, Bunn HF, Gallop PM (1977) J Clin Endocrinol Metab 44:859–864

    Article  CAS  Google Scholar 

  16. Larsen ML, Horder M, Mogensen EF (1990) N Engl J Med 323:1021–1025

    Article  CAS  Google Scholar 

  17. Lapolla A, Fedele D, Martano L, Arico' NC, Garbeglio M, Traldi P, Seraglia R, Favretto D (2001) J Mass Spectrom 36:370–378

    Article  CAS  Google Scholar 

  18. Stefanowicz P, Boratynski J, Kanska U, Petry I, Szewczuk Z (2001) Acta Biochim Pol 48:1137–1141

    CAS  Google Scholar 

  19. Yeboah FK, Yaylayan VA (2001) Nahrung 45:164–171

    Article  CAS  Google Scholar 

  20. Brock JW, Hinton DJ, Cotham WE, Metz TO, Thorpe SR, Baynes JW, Ames JM (2003) J Proteome Res 2:506–513

    Article  CAS  Google Scholar 

  21. Lapolla A, Fedele D, Reitano R, Arico NC, Seraglia R, Traldi P, Marotta E, Tonani R (2004) J Am Soc Mass Spectrom 15:496–509

    Article  CAS  Google Scholar 

  22. Mann M, Jensen ON (2003) Nat Biotechnol 21:255–261

    Article  CAS  Google Scholar 

  23. Kislinger T, Humeny A, Seeber S, Becker C-M, Pischetsrieder M (2002) Eur Food Res Technol 215:65–77

    Article  CAS  Google Scholar 

  24. Frolov A, Hoffmann P, Hoffmann R (2006) J Mass Spectrom 41:1459–1469

    Article  CAS  Google Scholar 

  25. Yaylayan V, Sporns P (1987) Food Chem 26:283–305

    Article  CAS  Google Scholar 

  26. Yaylayan V, Sporns P (1989) J Agric Food Chem 37:978–981

    Article  CAS  Google Scholar 

  27. Frolov A, Hoffmann R (2008) Ann N Y Acad Sci 1126:253–256

    Article  CAS  Google Scholar 

  28. Poduslo JF (1981) Anal Biochem 114:131–139

    Article  CAS  Google Scholar 

  29. Xiao-Chuan L (2006) Chin J Chromatogr 24:73–80

    Article  Google Scholar 

  30. Yue DK, McLennan S, Church DB, Turtle JR (1982) Diabetes 31:701–705

    Article  CAS  Google Scholar 

  31. Zhang Q, Tang N, Brock JW, Mottaz HM, Ames JM, Baynes JW, Smith RD, Metz TO (2007) J Proteome Res 6:2323–2330

    Article  CAS  Google Scholar 

  32. Klenk DC, Hermanson GT, Krohn RI, Fujimoto EK, Mallia AK, Smith PK, England JD, Wiedmeyer HM, Little RR, Goldstein DE (1982) Clin Chem 28:2088–2094

    CAS  Google Scholar 

  33. Zhang Q, Tang N, Schepmoes AA, Phillips LS, Smith RD, Metz TO (2008) J Proteome Res 7:2025–2032

    Article  CAS  Google Scholar 

  34. Lee B-S, Krishnanchettiar S, Lteef SS, Gupta S (2007) Int J Mass Spectrom 260:67–74

    Article  CAS  Google Scholar 

  35. Gadgil HS, Bondarenko PV, Treuheit MJ, Ren D (2007) Anal Chem 79:5991–5999

    Article  CAS  Google Scholar 

  36. Iberg N, Fluckiger R (1986) J Biol Chem 261:13542–13545

    CAS  Google Scholar 

  37. Gil H, Salcedo D, Romero R (2005) J Phys Org Chem 18:183–186

    Article  CAS  Google Scholar 

  38. Scaman C, Nakai S, Aminlari M (2006) Food Chem 99:368–380

    Article  CAS  Google Scholar 

  39. Walton DJ, Shilton BH (1991) Amino Acids 1:199–203

    Article  CAS  Google Scholar 

Download references

Acknowledgements

We thank Dr. Bernd Gründig for providing blood samples and Nicole Herth and Dr. Daniela Volke for preparation of the plasma samples. Financial support by the Deutsche Forschungsgemeinschaft (DFG, Graduiertenkolleg 378), the Free State Saxony and a scholarship of the DFG to A.F. are gratefully acknowledged.

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Author notes

  1. Andrej Frolov

    Present address: Department of Secondary Metabolism, Leibniz-Institute of Plant Biochemistry, Weinberg 3, 06120, Halle (Saale), Germany

Authors and Affiliations

  1. Institute of Bioanalytical Chemistry, Center for Biotechnology and Biomedicine, Faculty of Chemistry and Mineralogy, Universität Leipzig, Deutscher Platz 5, 04103, Leipzig, Germany

    Andrej Frolov & Ralf Hoffmann

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  1. Andrej Frolov
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  2. Ralf Hoffmann
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Correspondence to Ralf Hoffmann.

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Frolov, A., Hoffmann, R. Identification and relative quantification of specific glycation sites in human serum albumin. Anal Bioanal Chem 397, 2349–2356 (2010). https://doi.org/10.1007/s00216-010-3810-9

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  • DOI: https://doi.org/10.1007/s00216-010-3810-9

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