TRANSFERRED NUCLEAR OVERHAUSER ENHANCEMENT EXPERIMENTS SHOW THAT THE MONOCLONAL-ANTIBODY STREP-9 SELECTS A LOCAL MINIMUM CONFORMATION OF A STREPTOCOCCUS GROUP-A TRISACCHARIDE-HAPTEN

被引:34
作者
WEIMAR, T
HARRIS, SL
PITNER, JB
BOCK, K
PINTO, BM
机构
[1] SIMON FRASER UNIV, DEPT CHEM, BURNABY, BC V5A 1S6, CANADA
[2] BECTON DICKINSON RES CTR, RES TRIANGLE PK, NC 27709 USA
[3] CARLSBERG LAB, DEPT CHEM, DK-2500 VALBY, DENMARK
关键词
D O I
10.1021/bi00041a049
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Transferred nuclear Overhauser enhancement (TRNOE) experiments have been performed to investigate the bound conformation of the trisaccharide repeating unit of the Streptococcus Group A cell-wall polysaccharide. Thus, the conformations of propyl 3-O-(2-acetamido-2-deoxy-beta-D-glucopyranosyl 2-O-(alpha-L-rhamnopyranosyl)-alpha-L-rhamnopyranoside [C(A')B] (1) as a free ligand and when complexed to the monoclonal antibody Strep 9 were examined. Improved insights about the conformational preferences of the glycosidic linkages of the trisaccharide ligand showed that the free ligand populates various conformations in aqueous solution, thus displaying relatively flexible behavior. The NOE HNAc-H2A', which was not detected in previous work, accounts for a conformation at the beta-(1-->3) linkage with a phi angle of similar to 180 degrees. Observed TRNOEs for the complex are weak, and their analysis was further complicated by spin diffusion. With the use of transferred rotating-frame Overhauser enhancement (TRROE) experiments, the amount of spin diffusion was assessed experimentally, proving that ail of the observed long-range TRNOEs arose through spin diffusion. Four interglycosidic distances, derived from the remaining TRNOEs and TRROEs, together with repulsive constraints, derived from the absence of TRROE effects, were used as input parameters in simulated annealing and molecular mechanics calculations to determine the bound conformation of the trisaccharide. Complexation by the antibody results in the selection of one defined conformation of the carbohydrate hapten. This bound conformation, which is a local energy minimum on the energy maps calculated for the trisaccharide ligand, shows only a change from a +gauche to a -gauche orientation at the psi angle of the alpha-(1-->2) linkage when compared to the global minimum conformation. The results infer that the bound conformation of the Streptococcus Group A cell-wall polysaccharide is different from its previously proposed solution structure (Kreis et al., 1995).
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页码:13672 / 13681
页数:10
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