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a-Crystallin, an abundant mammalian lens protein made up of two subunits (aA- and aB-crystallin), is involved in the maintenance of the optimal refractive index in the lens. The protein is implicated in the pathophysiology of a large number of retinal diseases including cataract, age-related macular degeneration, diabetic retinopathy, and uveitis. a-Crystallin belongs to the small heat shock protein (sHSP) family, forms large oligomeric structures, and functions as a molecular chaperone appearing very early during embryonic development. To gain mechanistic insight into the structural and functional role of a-crystallin and its alterations in various retinal diseases, it is important to study the interaction chemistry with its known partners. The hydrophobic sites in a-crystallin have been studied extensively using environmentally sensitive fluorescent probes such as 4,4'-dianilino-1,1'-binaphthyl-5,5'-disulfonic acid dipotassium salt (bis-ANS) that interacts with both subunits of a-cystallin in 1:1 stoichiometry at 37 [degrees]C and diminishes the chaperone-like activity of the protein. Furthermore, it has been shown that ATP plays a crucial role in the association of [alpha]-crystallin with substrate proteins. We use surface plasmon resonance (SPR) to monitor the interactions of immobilized oligomeric recombinant [alpha]A subunit of human a-crystallin protein with bis-ANS and ATP. We assess the thermodynamic parameters and kinetics of such interactions at various temperatures. Our results indicate that bis-ANS binds to [alpha]A-crystallin with higher affinity when compared with ATP, although both [alpha]A-crystallin and [alpha]B-crystallin display fast interaction kinetics. Key words: [alpha]A-crystallin, ATP, bis-ANS, surface plasmon resonance (SPR). La cristalline [alpha], une proteine presente en abondance dans le cristallin des mammiferes, est constituee de deux sous-unites ([alpha]A et [alpha]B) et joue un role dans le maintien d'un indice de refraction optimal a l'interieur du cristallin. La proteine est impliquee dans la pathophysiologie d'un grand nombre de maladies de la retine, notamment la cataracte, la degenerescence maculaire liee a l'age, la retinopathie diabetique et l'uveite. La cristalline [alpha] appartient a la famille des petites proteines de choc thermique (sHSP). Elle forme de grandes structures oligomeriques et fonctionne comme un chaperon moleculaire, apparaissant tres tot dans le developpement embryonnaire. Afin de mieux comprendre les aspects mecanistiques du role fonctionnel et structural de la cristalline [alpha] et de ses anomalies dans diverses maladies retiniennes, il importe d'etudier la chimie de ses interactions avec les partenaires qu'on lui connait. Nous avons etudie en detail les sites hydrophobes de la cristalline [alpha] a l'aide de sondes fluorescentes sensibles a l'environnement telles que le sel dipotassique de l'acide 4,4'-dianilino-1,1'-binaphtyl-5,5'-disulfonique (bis ANS). Celui-ci interagit avec les deux sous-unites de la cristalline a dans un rapport stoechiometrique de 1:1 a 37 [degrees]C et reduit l'activite de type chaperon de la proteine. De plus, nous avons montre que l'ATP joue un role crucial dans l'association de la cristalline-a avec ses substrats proteiques. Nous avons utilise la resonance plasmonique de surface (SPR) pour observer les interactions avec le bis ANS et l'ATP d'un oligomere recombinant immobilise de la sous-unite [alpha]A de la cristalline [alpha] humaine. Nous avons evalue les parametres thermodynamiques et la cinetique de ces interactions a differentes temperatures. Nos resultats indiquent que le bis ANS se lie a la cristalline [alpha]A avec une plus grande affinite comparativement a l'ATP, bien que les deux presentent une cinetique d'interaction rapide. [Traduit par la Redaction] Mots-cles: cristalline [alpha]A, ATP, bis ANS, resonance plasmonique de surface (SPR)., Introduction [alpha]-Crystallin is a predominant protein present in the eye lens and comprises up to 40% of the total lens proteins. (1,2) It is made up of two subunits, [alpha]A [...]

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