Bioluminescent Bacteria
Bioluminescent bacteria are mainly found in seawater, where the majority form symbiotic relationships with other marine organisms. Their hosts have special organs where the bacteria reside safely and are fed, providing the host with light for protection, hunting, and communication.
There are three main genera of bioluminescent bacteria: Photobacterium, Vibrio, and Photorhabdus. Each genus and the species within them have unique properties; for example, vibrio harveyi lives on its own while vibrio fischeri is a symbiont.
The species of the Photobacterium genus are typically gram-negative cells with unsheathed flagella for movement. Photobacterium Phosphoreum, one species, can grow in an anaerobic environment and releases a bright blue-green light. Furthermore, it can be utilized to determine the relative toxicity of substances. We will be working with the lux genes of Photobacterium Phosphoreum. .
Bacterial Lux System
The bacterial lux system is principally composed of five genes - LuxA, LuxB, LuxC, LuxD, and LuxE.
[Diagram with their expression in nature, ie, CDABE w/ caption: In their host bacteria, the lux genes are not sequentially ordered, with A and B flanked by C, D, and E.]
The products from these genes associate to create two enzymatic complexes, with LuxAB coding for luciferase itself and LuxCDE coding for a fatty acid reductase complex that provides the aldehyde substrate for the light-generating reaction. Catalyzed by luciferase, these aldehydes then react with FMNH2 and oxygen, emitting a photon and producing a fatty acid, FMN, and water. FMNH2 is then regenerated by a flavin reductase.
[Complex diagram w/ elaborating caption: The alpha subunit of luciferase drives enzymatic activity, while the beta subunit offers structural support and stabilizes the alpha subunit as it undergoes conformational changes.]
To allow for continuous light output, the fatty acid reductase complex recycles the fatty acid produced in the luciferase-catalyzed reaction and converts it to a usable aldehyde. LuxC, LuxD, and LuxE code for a synthetase, transferase, and reductase, respectively. Together, they associate into a complex consisting of four of each enzyme.
[Complex diagram w/ elaborating caption: The enzymes are present in equal proportions in the fully-formed complex. Substrates are recruited by the transferase and moved to a synthetase-reductase complex. These associated enzymes create a microenvironments that stabilize the reaction intermediates.]
Because the luminescent yield of the system is based on the function of these two enzymatic complexes, modifying the expression levels of their coding genes allows us to control the system’s output.