A Single Amino Acid Mutation in Zebrafish (Danio rerio) Liver Bile Acid-binding Protein Can Change the Stoichiometry of Ligand Binding
Articolo
Data di Pubblicazione:
2007
Citazione:
A Single Amino Acid Mutation in Zebrafish (Danio rerio) Liver Bile Acid-binding Protein Can Change the Stoichiometry of Ligand Binding / S. Capaldi, M. Guariento, G. Saccomani, D. Fessas, M. Perduca, H.L. Monaco. - In: THE JOURNAL OF BIOLOGICAL CHEMISTRY. - ISSN 0021-9258. - 282:42(2007), pp. 31008-31018.
Abstract:
In all of the liver bile acid-binding proteins (L-BABPs) studied
so far, it has been found that the stoichiometry of binding is of
two cholate molecules per internal binding site. In this paper, we
describe the expression, purification, crystallization, and threedimensional
structure determination of zebrafish (Danio rerio)
L-BABP to 1.5A˚ resolution, which is currently the highest available
for a protein of this family. Since we have found that in
zebrafish, the stoichiometry of binding in the protein cavity is of
only one cholate molecule per wild type L-BABP, we examined
the role of two crucial amino acids present in the binding site.
Using site-directed mutagenesis, we have prepared, crystallized,
and determined the three-dimensional structure of co-crystals
of two mutants. The mutant G55R has the same stoichiometry of
binding as the wild type protein, whereas the C91T mutant
changes the stoichiometry of binding from one to two ligand
molecules in the cavity and therefore appears to be more similar
to the other members of the L-BABP family. Based on the presence
or absence of a single disulfide bridge, it can be postulated
that fish should bind a single cholate molecule, whereas
amphibians and higher vertebrates should bind two. Isothermal
titration calorimetry has also revealed the presence in the wild
type protein and the G55R mutant of an additional binding site,
different from the first and probably located on the surface of
the molecule.
Tipologia IRIS:
01 - Articolo su periodico
Elenco autori:
S. Capaldi, M. Guariento, G. Saccomani, D. Fessas, M. Perduca, H.L. Monaco
Link alla scheda completa: