This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Oligosaccharide was liberated by the method of Byers et al. (Byers HL et al. (1999) Glycobiology 9:469-479). Briefly, material was suspended in 0.1ml anhydrous hydrazine and incubated 5 hours at 95 C. Hydrazine was then evaporated under a gentle stream of dry air. Material was subsequently re-N acetylated by resuspending in 0.2ml saturated sodium bicarbonate, adding 10 l of acetic anhydride and incubating 1 hour at room temperature. The reaction mixture was then desalted on a 1ml Dowex H+ column and lyophilized. Oligosaccharides were isolated by method of Shimizu et al. (Shimizu Y et al. (2001) Carbohy. Res. 332:381-388). Briefly, lyophilized material was resuspended in a mixture of butonal:ethanol:water 4:1:1:v:v and loaded on a 1ml column of Avicel PH-101 cellulose which was preequilibrated in the same solvent after wash with 50% ethanol. The column was then eluted with 30ml butanol:ethanol:water 4:1:1, 10ml of 50% ethanol and 10ml of water. Oligosaccharides were noted in only the 50% ethanol eluated which was partially dried under a stream of dry air and lyophilized. Material was suspended in 0.5M dihydroxybenzoic acid in methanol and subjected by MALDI-MS on an Applied Biosystems Voyages DE MS which was run in the positive mode. All masses were calibrated by angiotension and ACTH controls run immediately before the samples.
Showing the most recent 10 out of 104 publications
