Engineering of bacterial luciferase from Vibrio campbellii for bioreportor application

Name: Paweenapon Chunthaboon

Organization : Mahidol University. Faculty of Science

Email : paweenapon.chu@student.mahidol.ac.th

Name: Ruchanok Tinikul

Organization : Mahidol University. Faculty of Science

ThaSH: Luciferases

ThaSH: Catalysts

ThaSH: Chemical reactions

ThaSH: Oxidation-reduction reaction

ThaSH: Chemiluminescence

Abstract: Bacterial luciferase (Lux) is bioluminescence enzyme. Lux generates blue-green light (lambda max 490 nm) by catalyzing the oxidation of reduced flavin mononucleotide (FMNH2), long-chain aldehyde and oxygen to yield oxidized FMN, carboxylic acid and water. Lux is heteromeric enzyme composing of alpha- and beta-subunits. To use in a eukaryotic reporter system, alpha- and beta-subunits of Lux have been fused to express as a single protein. The fusion Lux previously exhibited advantage over the current firefly and renilla luciferases in term of low background signal generation. However, a relatively large size of fusion Lux of 77.8 kDa and low reaction quantum yield (10-16 percent) still hamper the broad applications. In this study, the truncated fusion Lux was constructed by trimming C-terminus part of beta-subunit to obtain the smaller size of Lux that has ability to catalyze light reaction. Three truncated Luxs of Lux535, Lux555 and Lux595 were successfully constructed and expressed in E. coli. The result showed that among truncated constructs, only the constructed Lux535 could expressed as soluble enzyme. However, Lux535 could not process light reaction. This suggests that the beta-subunit portion of enzyme structure is very crucial for enzyme folding and active conformation of alpha-subunit.

King Mongkut's University of Technology North Bangkok. Central Library

Address: BANGKOK

Email: library@kmutnb.ac.th

Created: 2020

Modified: 2023-12-15

Issued: 2023-12-15

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BibliograpyCitation : In the Chemical Society of Thailand under the Patronage of Professor Dr. HRH Princess Chulabhorn in association, Thammasat University Faculty of Science and Technology. The Pure and Applied Chemistry International Conference 2020 (PACCON 2020) (CB54-59). Bangkok : Thammasat University

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