Modulation of catalytic functionality of alkaline phosphatase induced by semiconductor quantum dots: evidence of substrate-mediated protection
2016
Ghosh, Debasmita | Roy, Chandra Nath | Mondal, Somrita | Kundu, Somashree | Maiti, Susmita | Bag, Prasanta Kumar | Saha, Abhijit
Enzymes provide the critical means by which almost all biological reactions are catalyzed in a controlled manner. Methods to harness and exploit their properties are of strong current interest to the growing field of biotechnology. In view of many unique physical and optical characteristics that are advantageous for studying enzyme activity at quantum dot (QD)–bioconjugate interfaces, we endeavored to explore the possible influence on conformation and catalytic functionality of alkaline phosphatase (ALP), a clinical marker enzyme, by conducting in vitro enzymatic activity assay and using molecular spectroscopic techniques such as UV-visible absorption, fluorescence, circular dichroism (CD) and Fourier transform infrared (FTIR) spectroscopy. The resulting experimental data were then analyzed in the context of the classical Michaelis–Menten model. The results show almost 60% and 30% decrease in enzyme activity on interaction with 10 nM Cys-CdTe or Cys-CdS QDs, respectively, suggesting inhibition dependence on particle nature. CD spectra measurements indicated that QDs induced a decrease of α-helical content and an increase of β-sheet structure in ALP, resulting in the loosening and unfolding of the protein skeleton. Interestingly, when QDs and substrate were added simultaneously to enzyme molecules, catalytic inhibition was significantly reduced indicating protection of enzyme structure by substrate molecules. Further, we demonstrate that protein-encapsulated QDs do not affect the catalytic functionality of enzyme molecules. The present study provides valuable information which may have significant ramifications in various biomedical applications, such as biosensing, drug delivery, cellular imaging, etc.
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