title: Analysis of homodimeric avian and human galectins by two methods based on fluorescence spectroscopy: different structural alterations upon oxidation and ligand binding.
abstract: Spectroscopic monitoring is applied to detect structural alterations for homodimeric adhesion/growth-regulatory galectins. Mammalian galectin-1 and the avian ortholog CG-1B, due to their distinct patterns of cysteine positioning, can undergo oxidation. When monitoring tryptophan fluorescence anisotropy comparatively, an indicator of structural changes affecting rotational diffusion, segmental motion and/or fluorescence life time, reductions are seen in both cases upon oxidation. The decrease was especially marked for the human protein, more than 2-fold compared to the avian lectin. Using this approach to analyze binding of lactose, equilibrium and kinetic binding constants of both proteins were similar. This result is corroborated by fluorescence correlation spectroscopy with labeled proteins. Of note, the diffusion constant of CG-1B increased by 5.6% in the presence of lactose, as has been seen for the human protein. When processing the other two homodimeric avian galectins (CG-1A, CG-2) accordingly it was revealed that sequence homology does not translate into identical behavior. The diffusion constant of CG-1A was not affected, a slight decrease (-3.8%) was observed for CG-2. Obviously, alterations induced by oxidation and responses to ligand binding are different between these closely related proteins. Methodologically, the two spectroscopic techniques are proven to be sensitive and robust sensors for detecting intergalectin differences.
authors.affiliation: Department of Biotechnology and Biophysics, Julius-Maximilians University, Am Hubland, Biozentrum, 97074 Wuerzburg, Germany.
authors.forename: Antonia
authors.initials: A
authors.lastname: Göhler
authors.forename: Claudia
authors.initials: C
authors.lastname: Büchner
authors.forename: Sabine
authors.initials: S
authors.lastname: André
authors.lastname: Sören Doose
authors.forename: Herbert
authors.initials: H
authors.lastname: Kaltner
authors.forename: Hans-Joachim
authors.initials: HJ
authors.lastname: Gabius
mesh_terms: D000465:Algorithms; D000818:Animals; D030161:Avian Proteins / Q000737:chemistry / Q000378:metabolism*; D002645:Chickens; D019327:Copper Sulfate / Q000378:metabolism / Q000494:pharmacology; D004058:Diffusion / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D005454:Fluorescence Polarization / Q000379:methods*; D037483:Galectin 1 / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D007785:Lactose / Q000378:metabolism / Q000494:pharmacology; D008024:Ligands; D010084:Oxidation-Reduction; D011485:Protein Binding; D055503:Protein Multimerization; D013050:Spectrometry, Fluorescence / Q000379:methods*; D013329:Structure-Activity Relationship
chemical_list: D030161:Avian Proteins; D037483:Galectin 1; D008024:Ligands; D007785:Lactose; D019327:Copper Sulfate