Journal
MATERIALS SCIENCE AND ENGINEERING B-ADVANCED FUNCTIONAL SOLID-STATE MATERIALS
Volume 202, Issue -, Pages 46-53Publisher
ELSEVIER
DOI: 10.1016/j.mseb.2015.09.006
Keywords
Trypsin; Gold nanoparticle; Stoichiometry; Morphological change; Enhanced activity
Funding
- Amity University Haryana (AUH)
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Trypsin (Trp) is arguably the most important member of the serine proteases. Constructs made up of gold nanoparticles (GNP) with trypsin have been known to exhibit increased efficiency and stability in various experiments. Here we report simple Trp-GNP constructs mixed in different trypsin-to-GNP ratios which exhibit higher efficiencies in biochemical assay, varying resistance to autolysis and higher ability in cell trypsinization. Trp-GNP constructs in different trypsin-to-GNP ratios exhibit prolonged and sustained activity compared to native trypsin in N-alpha-p-benzoyl-p-nitroanilide (BAPNA) assay as monitored by UV-Visible spectroscopy. The activity was monitored as a function of decreasing rate of linear release of p-nitro aniline (resulting from the cleavage of BAPNA by trypsin) with time during the assay, whose absorbance was measured at 410nm (lambda(max) p-nitro aniline). We have done extensive studies to understand structural basis of this trypsin GNP interaction by using atomic force microscopy (AFM), transmission electron microscopy (TEM) and circular dichroism (CD) techniques. Our findings suggest that on interaction, the gold nanoparticles probably form an adherent layer on trypsin that effectively changes the morphology and dimensions of the nanoconstructs. However, trypsin-to-GNP ratio is extremely important, as higher concentration of GNP might damage the conformation of protein. Stability studies related to denaturation show that 1:1 Trp-GNP constructs exhibit maximum stability and high efficiency in all assays performed. (C) 2015 Elsevier B.V. All rights reserved.
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