Tanaya Chatterjee1, Barun K Chatterjee2, Tultul Saha3, Kazi Mirajul Hoque3, Pinak Chakrabarti4. 1. Department of Biochemistry, Bose Institute, P1/12 CIT Scheme VIIM, Kolkata 700054, India. Electronic address: tanaya@jcbose.ac.in. 2. Department of Physics, Bose Institute, 93/1, APC Road, Kolkata 700009, India. 3. Division of Molecular Pathophysiology, National Institute of Cholera & Enteric Diseases, P-33, CIT Road, Scheme XM, Beliaghata, Kolkata 700010, India. 4. Department of Biochemistry, Bose Institute, P1/12 CIT Scheme VIIM, Kolkata 700054, India.
Abstract
BACKGROUND: Accessory cholera enterotoxin (Ace) is a classical enterotoxin produced by Vibrio cholerae, the causative agent for cholera. Considering the crucial role of Ace in pathogenesis of cholera, we explored the modulation of structure/function of Ace using gold nanoparticles (AuNPs) of different size and shape - spherical (AuNS10 and AuNS100, the number indicating the diameter in nm) and rod (AuNR10). METHODS: Biophysical techniques have been used to find out structural modulation of Ace by AuNPs. Effect of AuNP on Ace conformation was monitored by far-UV CD; urea-induced unfolding and binding of Ace to various AuNPs were studied by tryptophan fluorescence. In vivo experiments using mouse ileal loop and Ussing chamber were carried out to corroborate biophysical data. RESULTS: Biophysical data revealed degradation of Ace by AuNR10 and AuNS100, not by AuNS10. The feature of AuNR10 having high aspect ratio, but with the same transverse diameter as that of AuNS10 enabled us to explore the importance of morphology on modulation of protein structure/function. The equilibration time for adsorption shows dependence on the radius of curvature, being largest for AuNR10. In vivo experiments revealed the efficacy of AuNR10 and AuNS100 for reduced fluid accumulation, indicative of the loss of activity of Ace. CONCLUSIONS: We show how biophysical studies and in vivo experiments go hand-in-hand in establishing the efficacy and role of size/shape of AuNPs on a toxin structure. GENERAL SIGNIFICANCE: The effect of AuNP on toxin depends on its morphology. The targeted modulation of Ace could be of therapeutic benefit for gastrointestinal disorders.
BACKGROUND: Accessory cholera enterotoxin (Ace) is a classical enterotoxin produced by Vibrio cholerae, the causative agent for cholera. Considering the crucial role of Ace in pathogenesis of cholera, we explored the modulation of structure/function of Ace using gold nanoparticles (AuNPs) of different size and shape - spherical (AuNS10 and AuNS100, the number indicating the diameter in nm) and rod (AuNR10). METHODS: Biophysical techniques have been used to find out structural modulation of Ace by AuNPs. Effect of AuNP on Ace conformation was monitored by far-UV CD; urea-induced unfolding and binding of Ace to various AuNPs were studied by tryptophan fluorescence. In vivo experiments using mouse ileal loop and Ussing chamber were carried out to corroborate biophysical data. RESULTS: Biophysical data revealed degradation of Ace by AuNR10 and AuNS100, not by AuNS10. The feature of AuNR10 having high aspect ratio, but with the same transverse diameter as that of AuNS10 enabled us to explore the importance of morphology on modulation of protein structure/function. The equilibration time for adsorption shows dependence on the radius of curvature, being largest for AuNR10. In vivo experiments revealed the efficacy of AuNR10 and AuNS100 for reduced fluid accumulation, indicative of the loss of activity of Ace. CONCLUSIONS: We show how biophysical studies and in vivo experiments go hand-in-hand in establishing the efficacy and role of size/shape of AuNPs on a toxin structure. GENERAL SIGNIFICANCE: The effect of AuNP on toxin depends on its morphology. The targeted modulation of Ace could be of therapeutic benefit for gastrointestinal disorders.