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Glycobiology, Vol 8, 1007-1012, Copyright © 1998 by Society for Glycobiology
V Sharma, VR Srinivas, P Adhikari, M Vijayan and A Surolia
The ability to discriminate between galactose and N- acetylgalactosamine,
observed in some lectins, is crucial for their biological activity as well
as their usefulness as tools in biology and medicine. However, the
molecular basis of differential binding of lectins to these two sugars is
poorly understood. Peanut agglutinin (PNA) is one of the few
galactose-specific legume lectins which does not bind N-
acetylgalactosamine at all and is, therefore, ideal for the study of the
basis of specificity towards C-2 substituted derivatives of
galactopyranosides. Examination of the three-dimensional structure of PNA
in complex with lactose revealed the presence of both a longer loop and
bulkier residues in the region surrounding the C-2 hydroxyl of the
galactopyranoside ring, which can sterically prevent the accommodation of a
bulky substituent in this position. One such residue, is a glutamic acid at
position 129 which protrudes into the binding site and perhaps directly
obstructs any substitution at the C-2 position. Two mutants in bacterially
expressed PNA were therefore constructed. These were E129D and E129A, in
which Glu129 was replaced by Asp and Ala, respectively. The specificity of
the mutants for galactose, galactosamine, and N- acetylgalactosamine was
examined through observing the inhibition of hemagglutination and binding
of the lectin to immobilized asialofetuin. The results showed that the
affinity of E129A and E129D for C-2-substituted derivatives of the
galactose varies. The mutant E129D showed significant binding towards N-
acetylgalactosamine, suggesting that the residue Glu 129 is crucial in
imparting exclusive galactose-specificity upon PNA. This study not only
attempts to provide an explanation for the inability of PNA to accommodate
C-2-substituted derivatives at its primary subsite, but also seeks to
present a basis for engineering lectins with altered specificities.
ORIGINAL ARTICLES
Molecular basis of recognition by Gal/GalNAc specific legume lectins: influence of Glu 129 on the specificity of peanut agglutinin (PNA) towards C2-substituents of galactose
Molecular Biophysics Unit, Indian Institute of Science, Bangalore 560 012, India.
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