Evaluation of the Site Specific Protein Glycation and Antioxidant Capacity of Rare Sugar−Protein/Peptide Conjugates

Protein−sugar conjugates generated in nonenzymatic glycation of α-lactalbumin (LA) with rare sugars [d-allose (All) and d-psicose (Psi)] and alimentary sugars as controls [d-glucose (Glc) and d-fructose (Fru)] were qualitatively determined by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS). Mass spectra revealed that the extent of glycation at lysine residues on LA with d-aldose molecules was very much higher than that of glycation with d-ketose molecules. To identify the specific site of glycation, the peptide mapping was established from protease V8 digestion, using a combination of computational cutting of proteins and MALDI-TOF-MS. As compared to peptide mapping, three and seven glycation sites were located in the primary structure of LA−ketose and LA−aldose conjugates, respectively. On the other hand, the antioxidant activities of protein−sugar conjugates and their peptic hydrolysates were investigated by 1,1-diphenyl-2-picrylhydrazyl radical scavenging method. The antioxidant activities of proteins/peptides glycated with rare sugars were significantly higher than those modified with the control sugars. The results indicated that the glycation degree and position were not markedly different between rare sugar and corresponding control sugar, but the antioxidant properties of protein and its hydrolysate were significantly enhanced by modifying with rare sugar. Keywords: α-Lactalbumin; rare sugar; glycation; antioxidant activity; peptide-mapping; MALDI-TOF-MS