| Project/Area Number |
06680573
|
|---|
| Research Category |
Grant-in-Aid for General Scientific Research (C)
|
| Allocation Type | Single-year Grants |
|---|
| Research Field |
Structural biochemistry
|
|---|
| Research Institution | GUNMA UNIVERSITY |
|---|
Principal Investigator |
KOCHIBE Naohisa DEPARTMENT OF BIOLOGY,FACULTY OF EDUCATION PROFESSOR, 教育学部, 教授 (60008266)
|
|---|
| Co-Investigator(Kenkyū-buntansha) |
YAZAWA Shin DEPARTMENT OF LEGAL MEDICINE,SCHOOL OF MEDICINE INSTRUCTOR, 医学部, 講師 (10008386)
|
|---|
| Project Period (FY) |
1994 – 1995
|
| Project Status |
Completed (Fiscal Year 1995)
|
| Budget Amount *help |
¥2,200,000 (Direct Cost: ¥2,200,000)
Fiscal Year 1995: ¥500,000 (Direct Cost: ¥500,000)
Fiscal Year 1994: ¥1,700,000 (Direct Cost: ¥1,700,000)
|
|---|
| Keywords | microbead / lectin / sugar chain / glycopepetide / agglutination / 固定化糖鎖物質 / ミクロスフィア / 結合特異性 / 複合糖鎖 / 複合糖質 |
|---|
| Research Abstract |
A simple and reliable method for detection of and analyzing lectin activities is developed by using microbeads on which sugar chains had been immobilized. For convenience in handling and adjusting lectin activity, the lectin used as a counterpart was also immobilized to the microbeads.
Amino group on the surface of Honensphere (2.5 mum in diameter) was succinylated with succinic anhydride, and then converted to active ester with cardodiimide. Amino group in the glycopeptide and lectin molecules was coupled to the active ester in the presence of N-hydroxysuccinimide under mild alkaline condition to give "sugar-bead" and "lectin-bead", respectively. The glycopeptides were derived from mucin type and Asn-linked type glycoprotems of which sugar structure has bead well known. Each of twelve lectins of known and 2 novel lectins was coupled to the bead.
Results of agglutination reaction of sugar-bead and lectin-bead showed good agreement with the reported lectin-sugar structure relationships, indicating that the present method is reliable. When a sugar-bead was sequentially treated with exoglycosidase, the lectin-bead accessible to the sugar-bead changed at every step of treatment, i.e. each lectin-bead reacted to the exposed sugar moiety. This result indicates that the binding properties could be analyzed by using the sugar-bead and its enzymetreated one. Indeed, two novel lectins from fruiting bodies of Basidiomycetes and Ascomycetes bound to a T antigen structure and strictly required a conformation of GalNAc at the reducing terminal.
Purification and properties of these lectins will be reported elsewhere.
Lectin-beads were applicable to examine cell surface sugar structures. Although it is only qualitative, the method is simple and reliable as compared to flow cytometry or fluorescence microscopy.
|
