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Implication of an unfavorable residue (Thr346) in intrinsic flexibility of firefly luciferase

Paper ID Volume ID Publish Year Pages File Format Full-Text
17102 42641 2012 7 PDF Available
Title
Implication of an unfavorable residue (Thr346) in intrinsic flexibility of firefly luciferase
Abstract

In order to better understand the functional role of an unusual residue (Thr346) of firefly luciferase mutagenesis at this residue was performed. Firefly luciferase, catalyzes the bioluminescence reaction and is an excellent tool as a reporter in nano-system biology studies. Nonetheless, the enzyme rapidly loses its activity at temperatures above 30 °C and this leads to reduced sensitivity and precision in analytical applications. Residue Thr346 in a connecting loop (341–348) of firefly luciferase is located in a disallowed region of Ramachandran plot. In this study, we have substituted this residue (T346) with anomalous dihedral angles with Val, Gly and Pro to clarify the role of this residue in structure and function of the enzyme using site-directed mutagenesis. Substitution of this unfavorable residue (T346) with atypical dihedral angles (ψ, φ) with other residues brought about an increase of thermostability and decrease of specific activity. Structural and functional properties of the mutants were analyzed using different spectroscopic methods. It seems that this residue is a critically conserved residue to support the functional flexibility for a fast kinetic bioluminescence reaction at the expense of lower stability.

► Thr346 of luciferase is located in a disallowed region of Ramachandran plot. ► T346 is substituted with Val, Gly and Pro. ► Substitution increased thermostability and decreased specific activity. ► This residue is a conserved residue to support functional flexibility of luciferase.

Keywords
Luciferase; Flexibility; Dihedral angles; Stability; Bioluminescence
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Implication of an unfavorable residue (Thr346) in intrinsic flexibility of firefly luciferase
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Publisher
Database: Elsevier - ScienceDirect
Journal: Enzyme and Microbial Technology - Volume 51, Issue 4, 10 September 2012, Pages 186–192
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
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Full-text PDF Download
Online Support
Any Questions? feel free to contact us