Bioluminescence and Applications

**3**

**Chapter 1**

*and Dan Close*

**Abstract**

Biotechnological Advances in

*Andrew Kirkpatrick, Tingting Xu, Steven Ripp, Gary Sayler* 

This chapter explores the history of the bioengineering advances that have been applied to common luciferase enzymes and the improvements that have been accomplished by this work. The primary focus is placed on firefly luciferase (FLuc), *Gaussia* luciferase (GLuc), *Renilla* luciferase (RLuc), *Oplophorus* luciferase (OLuc; NanoLuc), and bacterial luciferase (Lux). Beginning with the cloning and exogenous expression of each enzyme, their step-wise modifications are presented and the new capabilities endowed by each incremental advancement are highlighted. Using the historical basis of this information, the chapter concludes with a prospective on the overall impact these advances have had on scientific research and

provides an outlook on what capabilities future advances could unlock.

**Keywords:** firefly luciferase (FLuc), *Gaussia* luciferase (GLuc), *Renilla* luciferase (RLuc), *Oplophorus* luciferase (OLuc; NanoLuc),

**1.1 Historical perspective on the discovery of luciferase enzymes**

The bioluminescent phenotype, which is spread across a variety of different insects, bacteria, fungi, and marine animals, has intrigued mankind since before the dawn of the modern scientific era [1]. The discovery that proteins, which would come to be known as luciferases, were responsible for bioluminescent production can be traced to early experiments by Raphael Dubois, who was able to produce bioluminescence *in situ* by mixing the contents of click beetle abdomens in cold water and extracting the components required for light production [2]. However, it was not until the late 1940s that the first luciferase protein was successfully purified from fireflies [3]. Around that same time, bacterial luciferase was elucidated and successfully expressed *in situ* [4]. However, despite the progress made with these luciferases, it would be some time until biotechnology had advanced to the point where the genes responsible for their expression could be cloned and exogenously expressed, setting off the use of luciferases as tools for scientific discovery [5, 6]. Following the exogenous expression of the previously described firefly and bacterial luciferases, *Renilla* luciferase was isolated from the sea pansy *Renilla reniformis* [7] and *Oplophorus* luciferase was isolated from the deep-sea shrimp, *Oplophorus gracilirostris* [8]. Shortly thereafter, firefly luciferase was successfully

bacterial luciferase (Lux), biotechnology

**1. Introduction**

Luciferase Enzymes

## **Chapter 1**
