Amino Acids

, Volume 48, Issue 7, pp 1631–1639 | Cite as

The glycation of fibronectin by glycolaldehyde and methylglyoxal as a model for aging in Bruch’s membrane

  • Mai T. Thao
  • Elizabeth R. GaillardEmail author
Original Article


The purpose of the study is to identify the sites of modification when fibronectin reacts with glycolaldehyde or methylglyoxal as a model system for aging of Bruch’s membrane. A synthetic peptide consisting of the α5β1 integrin binding region of fibronectin was incubated with glycolaldehyde for 12 h or with methylglyoxal for 1 h at 37 °C. After tryptic digestion, the samples were analyzed with liquid chromatography–mass spectrometry (LC/MS). Tandem MS was used to determine the sites of modification. The adducts, aldoamine and N ε-carboxymethyl-lysine, attached preferably at lysine residues when the fibronectin peptide reacted with glycolaldehyde. When the fibronectin peptide reacted with methylglyoxal, modifications occurred at lysine and arginine residues. At lysine residues, N ε-carboxyethyl-lysine adducts were present. At arginine residues, hydroimidazolone and tetrapyrimidine adducts were present. Several advanced glycation endproducts were generated when fibronectin was glycated via glycolaldehyde and methylglyoxal. These results can help explain the structural changes Bruch’s membrane undergoes during aging.


Bruch’s membrane Fibronectin Retinal pigment epithelium Glycation Aging 


Compliance with ethical standards

Conflict of interests

The authors do not have competing interests and/or commercial relationships.

The work reported here does not invovle human participants or animal subjects.


  1. Adrover M, Vilanova B, Munoz F, Donoso J (2008) Kinetic study of the reaction of glycolaldehyde with two glycation target models. Ann N Y Acad Sci 1126:235–240CrossRefPubMedGoogle Scholar
  2. Ahmed N, Thornalley PJ, Dawczynski J, Franke S, Strobel J, Stein G, Haik GM (2003) Methylglyoxal-derived hydroimidazolone advanced glycation end-products of human lens proteins. Invest Ophthalmol Vis Sci 44(12):5287–5292CrossRefPubMedGoogle Scholar
  3. Beattie JR, Pawlak AM, Boulton ME, Zhang J, Monnier VM, McGarvey JJ, Stitt AW (2010) Multiplex analysis of age-related protein and lipid modifications in human Bruch’s membrane. FASEB J 24(12):4816–4824CrossRefPubMedPubMedCentralGoogle Scholar
  4. Brock JW, Cotham WE, Thorpe SR, Baynes JW, Ames JM (2007) Detection and identification of arginine modifications on methylglyoxal-modified ribonuclease by mass spectrometric analysis. J Mass Spectrom 42(1):89–100CrossRefPubMedGoogle Scholar
  5. Dobler D, Ahmed N, Song L, Eboigbodin KE, Thornalley PJ (2006) Increased dicarbonyl metabolism in endothelial cells in hyperglycemia induces anoikis and impairs angiogenesis by RGD and GFOGER motif modification. Diabetes 55(7):1961–1969CrossRefPubMedGoogle Scholar
  6. Duran-Jimenez B, Dobler D, Moffatt S, Rabbani N, Streuli CH, Thornalley PJ, Tomlinson DR, Gardiner NJ (2009) Advanced glycation end products in extracellular matrix proteins contribute to the failure of sensory nerve regeneration in diabetes. Diabetes 58(12):2893–2903CrossRefPubMedPubMedCentralGoogle Scholar
  7. Farboud B, Aotaki-Keen A, Miyata T, Hjelmeland LM, Handa JT (1999) Development of a polyclonal antibody with broad epitope specificity for advanced glycation endproducts and localization of these epitopes in Bruch’s membrane of the aging eye. Mol Vis 5:11PubMedGoogle Scholar
  8. Glenn JV, Beattie JR, Barrett L, Frizzell N, Thorpe SR, Boulton ME, McGarvey JJ, Stitt AW (2007) Confocal Raman microscopy can quantify advanced glycation end product (AGE) modifications in Bruch’s membrane leading to accurate, nondestructive prediction of ocular aging. FASEB J 21(13):3542–3552CrossRefPubMedGoogle Scholar
  9. Glenn JV, Mahaffy H, Wu K, Smith G, Nagai R, Simpson DA, Boulton ME, Stitt AW (2009) Advanced glycation end product (AGE) accumulation on Bruch’s membrane: links to age-related RPE dysfunction. Invest Ophthalmol Vis Sci 50(1):441–451CrossRefPubMedGoogle Scholar
  10. Glomb MA, Monnier VM (1995) Mechanism of protein modification by glyoxal and glycolaldehyde, reactive intermediates of the Maillard reaction. J Biol Chem 270(17):10017–10026CrossRefPubMedGoogle Scholar
  11. Handa JT, Verzijl N, Matsunaga H, Aotaki-Keen A, Lutty GA, te Koppele JM, Miyata T, Hjelmeland LM (1999) Increase in the advanced glycation end product pentosidine in Bruch’s membrane with age. Invest Ophthalmol Vis Sci 40(3):775–779PubMedGoogle Scholar
  12. Hasegawa G, Nakano K, Tsutsumi Y, Kondo M (1994) Effects of aldehyde-modified proteins on mesangial cell-matrix interaction. Diabetes Res Clin Pract 23(1):25–32CrossRefPubMedGoogle Scholar
  13. Hayashi CM, Nagai R, Miyazaki K, Hayase F, Araki T, Ono T, Horiuchi S (2002) Conversion of Amadori products of the Maillard reaction to N(epsilon)-(carboxymethyl)lysine by short-term heating: possible detection of artifacts by immunohistochemistry. Lab Invest 82(6):795–808CrossRefGoogle Scholar
  14. Howes KA, Liu Y, Dunaief JL, Milam A, Frederick JM, Marks A, Baehr W (2004) Receptor for advanced glycation end products and age-related macular degeneration. Invest Ophthalmol Vis Sci 45(10):3713–3720CrossRefPubMedGoogle Scholar
  15. Hussain AA, Starita C, Hodgetts A, Marshall J (2010) Macromolecular diffusion characteristics of ageing human Bruch’s membrane: implications for age-related macular degeneration (AMD). Exp Eye Res 90(6):703–710CrossRefPubMedGoogle Scholar
  16. Ida H, Ishibashi K, Reiser K, Hjelmeland LM, Handa JT (2004) Ultrastructural aging of the RPE-Bruch’s membrane-choriocapillaris complex in the d-galactose-treated mouse. Invest Ophthalmol Vis Sci 45(7):2348–2354CrossRefPubMedGoogle Scholar
  17. Lo TW, Westwood ME, McLellan AC, Selwood T, Thornalley PJ (1994) Binding and modification of proteins by methylglyoxal under physiological conditions. A kinetic and mechanistic study with N alpha-acetylarginine, N alpha-acetylcysteine, and N alpha-acetyllysine, and bovine serum albumin. J Biol Chem 269(51):32299–32305PubMedGoogle Scholar
  18. Murdaugh LS, Dillon J, Gaillard ER (2009) Modifications to the basement membrane protein laminin using glycolaldehyde and A2E: a model for aging in Bruch’s membrane. Exp Eye Res 89(2):187–192CrossRefPubMedGoogle Scholar
  19. Nagai R, Matsumoto K, Ling X, Suzuki H, Araki T, Horiuchi S (2000) Glycolaldehyde, a reactive intermediate for advanced glycation end products, plays an important role in the generation of an active ligand for the macrophage scavenger receptor. Diabetes 49(10):1714–1723CrossRefPubMedGoogle Scholar
  20. Nagaraj RH, Linetsky M, Stitt AW (2012) The pathogenic role of Maillard reaction in the aging eye. Amino Acids 42(4):1205–1220CrossRefPubMedGoogle Scholar
  21. Rabbani N, Thornalley PJ (2011) Glyoxalase in diabetes, obesity and related disorders. Semin Cell Dev Biol 22(3):309–317CrossRefPubMedGoogle Scholar
  22. Rabbani N, Thornalley PJ (2012) Methylglyoxal, glyoxalase 1 and the dicarbonyl proteome. Amino Acids 42(4):1133–1142CrossRefPubMedGoogle Scholar
  23. Schutt F, Bergmann M, Holz FG, Kopitz J (2003) Proteins modified by malondialdehyde, 4-hydroxynonenal, or advanced glycation end products in lipofuscin of human retinal pigment epithelium. Invest Ophthalmol Vis Sci 44(8):3663–3668CrossRefPubMedGoogle Scholar
  24. Thorpe SR, Baynes JW (2003) Maillard reaction products in tissue proteins: new products and new perspectives. Amino Acids 25(3–4):275–281CrossRefPubMedGoogle Scholar
  25. Ulrich P, Cerami A (2001) Protein glycation, diabetes, and aging. Recent Prog Horm Res 56:1–21CrossRefPubMedGoogle Scholar
  26. Wang Z, Paik DC, Del Priore LV, Burch RL, Gaillard ER (2005) Nitrite-modified extracellular matrix proteins deleteriously affect retinal pigment epithelial cell function and viability: a comparison study with nonenzymatic glycation mechanisms. Curr Eye Res 30(8):691–702CrossRefPubMedGoogle Scholar
  27. Yamada Y, Ishibashi K, Bhutto IA, Tian J, Lutty GA, Handa JT (2006) The expression of advanced glycation endproduct receptors in rpe cells associated with basal deposits in human maculas. Exp Eye Res 82(5):840–848CrossRefPubMedGoogle Scholar
  28. Zhang Q, Ames JM, Smith RD, Baynes JW, Metz TO (2009) A perspective on the Maillard reaction and the analysis of protein glycation by mass spectrometry: probing the pathogenesis of chronic disease. J Proteome Res 8(2):754–769CrossRefPubMedPubMedCentralGoogle Scholar
  29. Zhang Z, Yang Z, Zhu B, Hu J, Liew CW, Zhang Y, Leopold JA, Handy DE, Loscalzo J, Stanton RC (2012) Increasing glucose 6-phosphate dehydrogenase activity restores redox balance in vascular endothelial cells exposed to high glucose. PLoS ONE 7(11):e49128CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer-Verlag Wien 2016

Authors and Affiliations

  1. 1.Department of Chemistry and BiochemistryNorthern Illinois UniversityDeKalbUSA
  2. 2.Department of Biological SciencesNorthern Illinois UniversityDeKalbUSA

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