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Fluorescence spectroscopic studies on tryptophan at the saccharide‐binding site of castor bean hemagglutinin
Author(s) -
Yamasaki Nobuyuki,
Absar Nural,
Funatsu Gunki
Publication year - 1989
Publication title -
journal of molecular recognition
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.401
H-Index - 79
eISSN - 1099-1352
pISSN - 0952-3499
DOI - 10.1002/jmr.300010402
Subject(s) - tryptophan , fluorescence , chemistry , hemagglutinin (influenza) , binding site , biochemistry , amino acid , physics , quantum mechanics , gene
The environment of tryptophan in castor bean hemagglutinin (CBH) was analyzed fluorescence spectroscopy with regard to saccharide binding. Upon binding of specfic saccharides, the fluorescence maximum of 333 nm of CBH shifted to a wavelenght 2 nm shorter, owing to the change in the environment of tryptophan at the saccharide‐binding site. By analyzing the change in the fluorescence intensity at 320nm as a function of concentration of saccharides, the association constants for binding of saccharides to CBH were determined. The results suggest that the saccharide‐binding site on each B‐chain is actually composed of a subsite with which the saccharide residue linked to galactopyranoside at the non‐reducing end can interact, and another site which recognizes the galactopyranoside moiety. Quenching data indicated that five out of 22 tryptophans in CBH are surface‐localized and are available for quenching with both KI and acrylamide, and three other tryptophans are buried and are available only to acrylamide. Binding of raffinose to CBH decreased by 2 the number of tryptophan residues accesible to quenchers in the CBH molecule. We speculate that raffinose binds to CBH in such a manner as to shield the tryptophan located at the subsite from quenching by KI and acrylamide. The results also suggest that the tryptophan located at the subsite from quenching by KI and acrylamide. The results also suggest that the tryptophan residue atresidu at the saccharide‐binding site on each B‐chain is localized near the surface, and present in the positively charged environment.