Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy
Enhancement in chaperone activity of human αA-crystallin by nanochaperone gold nanoparticles: Multispectroscopic studies on their molecular interactions
Graphical abstract
Introduction
The bulk of the eye lens is made of firmly packed fibres cells, which are devoid of some major organelle. For proper vision, light entering in the lens through cornea is transmitted and focused on the retina. The lens fibre cells involved in the above process are mostly comprised of crystallin proteins. These proteins are present at a high concentration and account for the transparency of lens. Crystallins are structural proteins and categorized in to three types (α, β and γ). Alpha-crystallin possesses chaperone-like behaviour [1], [2], [3]. Alpha-crystallin contributes ∼ 28% to the total protein content of the lens and maintains the required refractive index. Two subunits of α-crystallin (αA and αB) exist in 3:1 proportion [4]. Under stressful conditions, partial unfolding of β and γ-crystallins cause their aggregation. As a molecular chaperone, α-crystallin forms stable complexes with partially unfolded β and γ-crytallins to suppress their aggregation [5]. However, with aging, the chaperone activity of α-crystallin deteriorates and accumulation of protein aggregates begins in the lens. In normal condition, the metal ions like Cu2+ and Zn2+ are kept bound by α-crystallin [6], which suppresses metal-mediated oxidative protein aggregation in the lens [7]. But complexation between α-crystallin and its substrates (β and γ-crystallins) in aged lens deprive the metal binding ability of α-crystallin [8]. In that situation, β/γ-crystallins start to form protein aggregates, which scatter light and increase the opacity of the lens [9], [10]. This is commonly referred as cataract and recognized as the main cause of blindness worldwide. The aggregates thus formed remain in the lens for lifetime and can only be removed surgically.
An alternate non-surgical approach can be developed by increasing the chaperone activity of α-crystallin in aged lens which would delay the formation of cataract. Attempts to enhance the chaperone activity of α-crystallin had been reported earlier using small molecules, antiglycating agents etc. [11]. Earlier our group had also reported a synthetic Schiff base molecule having positive cooperative effect on the chaperone activity of human αA-crystallin (HAA) [12]. Further, molecular interactions between HAA and that Schiff base were studied to find out the mode of action [13]. Application of nanoparticles in the augmentation of the chaperone activity of αA-crystallin would be a remarkable feat in prolonging the development of cataract. In the present work, gold nanoparticles (AuNps) were used to enhance the chaperone activity of HAA. Selection of AuNPs in this purpose was based on earlier works, which had reported the potency of AuNPs to serve as nano-chaperone [14], [15], [16], [17], [18], [19]. Moreover, AuNPs was emerged as potent inhibitor against aggregation of different proteins and peptides [20]. Recently, our group had also reported the inhibition of amyloid fibrillation of human γD-crystallin (HGD) by AuNPs [21]. Based on these reports, application of AuNPs as nano-chaperone in the presence of HAA during aggregation of HGD through partial unfolding was attempted. Furthermore, interactions between HAA and AuNPs were studied.
Section snippets
Materials and methods
The reagents and culture media were purchased from Himedia and SRL. Absorption spectra were recorded on Hitachi UH5300 and fluorescence spectra on Shimadzu RF-5301PC and RF-6000. All the experiments were carried out in phosphate buffer (10 mM, pH 7.0). Over-expression and purification of HGD and HAA was accomplished following the procedure described earlier by Chauhan et al. [12].
Gold nanoparticles (AuNPs) were prepared from HAuCl4 (1 mM) through reduction by using tri-sodium citrate (20 mM)
Results and discussion
Gold nanoparticles (AuNPs) have earlier been reported as nanochaperone [14], [15], [16], [17], [18], [19]. The present work was thought with an idea to boost up the chaperone activity of HAA under the limiting conditions in an aged lens. Therefore, the effect of nanochaperone like AuNPs on the chaperone activity of HAA was checked. To assess the chaperone activity of HAA, aggregation of its substrate (HGD) was initiated during the refolding of the protein, which was previously unfolded by
Conclusions
Multispectroscopic studies on the interactions between HAA and AuNPs were presented in this work. Formation of ground-state complex between HAA and nanoparticles leads to static quenching of Trp fluorescence. Thermodynamic parameters suggest spontaneous bioconjugation. Hydrogen bonding and van der Waals interactions were involved in the complexation between HAA and AuNPs. The Trp microenvironment of HAA was not altered by AuNPs. The nanoparticles also did not perturb the secondary structure of
Declaration of Competing Interest
The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
Acknowledgements
AS and KSG are thankful to the Director, NIT Hamirpur for providing research facilities. AS is obligated to NIT Hamirpur for Senior Research Fellowship. AS is thankful to Dr. Arvind Kumar and Dr. Tapas Palai (NIT Hamirpur), Prof. Pramit Kumar Chowdhury (IIT Delhi), Dr. Anjan Chakraborty and Avijit Maity (IIT Indore) for some instrumental supports.
References (35)
- et al.
The chaperone activity of bovine α-crystallin
J. Biol. Chem.
(1994) - et al.
α-Crystallin as a molecular chaperone
Prog. Retin. Eye Res.
(1999) - et al.
Partially folded aggregation intermediates of human γD-, γC-, and γS-crystallin are recognized and bound by human αB-crystallin chaperone
J. Mol. Biol.
(2010) - et al.
Effects of divalent metal ions on the αB-crystallin chaperone-like activity: spectroscopic evidence for a complex between copper (II) and protein
J. Inorg. Biochem.
(2004) - et al.
Lens aging: effects of crystallins
Biochim. Biophys. Acta
(2009) - et al.
Ageing and vision: structure, stability and function of lens crystallins
Prog. Biophys. Mol. Biol.
(2004) - et al.
Studies on molecular interactions between Schiff bases and eye lens chaperone human αA-crystallin
J. Lumin.
(2017) - et al.
Amyloid fibril formation by β-lactoglobulin is inhibited by gold nanoparticles
Int. J. Biol. Macromol.
(2014) - et al.
Inhibition of amyloid fibrillation of human γD-crystallin by gold nanoparticles: studies at molecular level
Spectrochim. Acta A
(2020) - et al.
Extinction coefficient of gold nanoparticles with different sizes and different capping ligands
Colloids Surf. B
(2007)