Detailed oligosaccharide structures of human integrin alpha 5 beta 1 analyzed by a three-dimensional mapping...

Structures of N-linked oligosaccharides obtained from human integrin alpha 5 beta 1 are described. Integrin alpha 5 beta 1 (4.5 mg) was purified from human placenta and digested using trypsin and chymotrypsin. N-linked oligosaccharides were released from the glycopeptides by digestion with glycoamidase A (from almond). The reducing ends of the oligosaccharides were derivatized with 2-aminopyridine. The pyridylamino-oligosaccharides were separated and these structures were identified by a three-dimensional HPLC mapping technique on three kinds of HPLC columns [Takahashi, N., Nakagawa, H., Fujikawa, K., Kawamura, Y. & Tomiya, N. (1995) Anal. Biochem. 226, 139-146]. Finally, 35 different oligosaccharide structures were identified, 10 of which were neutral, 6 mono-sialyl, 10 di-sialyl, 7 tri-sialyl and 2 tetra-sialyl. The molar ratio of neutral, mono-sialyl, di-sialyl, tri-sialyl and tetra-sialyl oligosaccharides was 20.8%, 24.8%, 27.7%, 18.1% and 8.6%, respectively. High-mannose-type oligosaccharides accounted for only 1.5% of the total. The remaining oligosaccharides were all complex type. The most predominant structure was the diantennary di-alpha-(2,3)-sialyl fucosyl. Major linking of sialic acid was alpha-(2,3)-linkage, and over 50% of all oligosaccharides were fucosylated at the N-acetylglucosamine residue of the reducing end.