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ISSN
1672-9145
Acta Biochim Biophys Sin
2005, 37(1): 47–54
CN 31-1940/Q
Effect of Amino Acid Residue and
Oligosaccharide Chain Chemical Modifications on Spectral and Hemagglutinating
Activity of Millettia dielsiana Harms. ex Diels. Lectin
Shun GAO, Jie AN, Chuan-Fang
WU, Ying GU, Fang CHEN, Yuan YU1, Qia-Qing WU1*, and Jin-Ku BAO*
College of Life Sciences,
Sichuan University, Chengdu 610064, China;
1Chengdu
Institute of Biology, Chinese Academy of Sciences, Chengdu 610041, China
Abstract The
effects of modifying the carbohydrate chain and amino acids on the conformation
and activity of Millettia dielsiana Harms. ex Diels. lectin (MDL) were
studied by hemagglutination, fluorescence and circular dichroism analysis. The
modification of tryptophan residues led to a compete loss of hemagglutinating
activity; however, the addition of mannose was able to prevent this loss of
activity. The results indicate that two tryptophan residues are involved in the
carbohydrate-binding site. Modifications of the carboxyl group residues
produced an 80% loss of activity, but the presence of mannose protected against
the modification. The results suggest that the carboxyl groups of aspartic and
glutamic acids are involved in the carbohydrate-binding site of the lectin.
However, oxidation of the carbohydrate chain and modification of the histidine
and arginine residues did not affect the hemagglutinating activity of MDL.
Fluorescence studies of MDL indicate that tryptophan residues are present in a
relatively hydrophobic region, and the binding of mannose to MDL could quench
tryptophan fluorescence without any change in lmax. The circular dichroism spectrum showed that all of these
modifications affected the conformation of the MDL molecule to different
extents, except the modification of arginine residues. Fluorescence quenching
showed that acrylamide and iodoacetic acids are able to quench 77% and 98% of
the fluorescence of tryptophan in MDL, respectively. However, KI produced a
barely perceptible effect on the fluorescence of MDL, even when the
concentration of I was 0.15 M. This demonstrates that most
of tryptophan residues are located in relatively hydrophobic or negatively
charged areas near the surface of the MDL molecule.
Key words Millettia
dielsiana Harms. ex Diels. lectin (MDL); chemical modification;
hemagglutinating activity; circular dichroism; fluorescence quenching
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Received: September 10, 2004 Accepted: December 10, 2004
This work was supported by a grant from the National Natural Science Foundation of China (No. 30000032)
*Corresponding authors:
Qia-Qing WU: Tel, 86-28-82900607; E-mail, [email protected]
Jin-Ku BAO: Tel, 86-28-85410672; E- mail, [email protected]