Expression and Purification of Firefly Luciferase and its Interaction with Cadmium Telluride Quantum Dot

Document Type: Article

Authors

1 Biotechnology, Science and Research Branch, Islamic Azad University, Tehran, Iran

2 Tarbiat Modares University

3 Department of Medical Nanotechnology, Applied Biophotonic Research center, Science and Research Branch, Islamic Azad University, Tehran, Iran

4 Co-funder & CEO Nora gene pishro,knowledge base company,Tehran, Iran

5 Miniistry of Health_Environmental and Occupational health center, Tehran, Iran Abstract

Abstract

Firefly luciferase is a monomeric enzyme of 62 kDa that catalyzes emission of green to yellow region, typically 550–570 nm upon reaction with d-luciferin, ATP, and molecular oxygen. Semiconductor nanocrystal, also known as the quantum dots (QDs), are nanoscaled inorganic particles in size range of 1-10nm. QDs have properties, such as sharp and symmetrical emission spectra, size-dependent emission, good chemical and photostability and high quantum yield. In this study, recombinant P. pyralis luciferase was expressd and then purified based on N -terminal His-tag. Then the effects of the Cadmium Tellurium QD(6.2nm) on the tertiary structure, kinetic properties, Bioluminescence Resonance Energy Transfer, thermal stability and remaining activity of luciferase was assayed using fluorescence spectroscopy and bioluminescence assay. The results showed that the CdTe QD affects the tertiary structure of the luciferase enzyme. The kinetic parameters of the enzyme also changed, as well as the thermal stability. The remaining activity was decrease in the presence of quantum dot compared to the native enzyme and BRET also was not observed in the presence of QD.

Keywords