Bioluminescence bacterial luciferases

Bioluminescence can also be used as the basis for immunoassay. For example, bacterial luciferase has been used in a co-immobilized system to detect and quantify progesterone using a competitive immunoassay format (34), and other luciferase-based immunoassays have been used to quantify insulin, digoxin, biotin, and other clinically important analytes (35). 

Balny, C., and Hastings, J. W. (1975). Fluorescence and bioluminescence of bacterial luciferase intermediates. Biochemistry 14 4719-4723. 

Kurfuerst, M., Macheroux, P., Ghisla, S., and Hastings, J. W. (1989). Bioluminescence emission of bacterial luciferase with 1 -deaza-FMN. Evidence for the noninvolvement of N(l)-protonated flavin species as emitters. Eur. J. Biochem. 181 453 157. 

Lee, J., O Kane, D. J., and Gibson, B. G. (1989). Bioluminescence spectral and fluorescence dynamics study of the interaction of lumazine protein with the intermediates of bacterial luciferase bioluminescence. Biochemistry 28 4263-4271. 

Legocki, R. P., Legocki, M., Baldwin, T. O., and Szalay, A. A. (1986). Bioluminescence in soybean root nodules demonstration of a general approach to assay gene expression in vivo by using bacterial luciferase. Proc. Natl. Acad. Sci. USA 83 9080-9084. 

Meighen, E. A., and Bartlet, I. (1980). Complementation of subunits from different bacterial luciferases. Evidence for the role of the (3 subunit in the bioluminescent mechanism. J. Biol. Chem. 255 11181— 11187. 

Sinclair, J. F., Waddle, J. J., Waddill, E. F., and Baldwin, T. O. (1993). Purified native subunits of bacterial luciferase are active in the bioluminescence reaction but fail to assemble into the a(3 structure. Biochemistry 32 5036-5044. 

Zeng, J., and Jewsbury, R. A. (1993). Enhanced bioluminescence of bacterial luciferase induced by metal ions and their complexes. In Szalay, A. A., et al. (eds.), Biolumin. Chemilumin., Proc. Int. Symp., 7th, pp. 173-177. Wiley, Chichester, UK. 

Bacterial bioluminescence, 30-46 factors required, 31 general scheme, 32 in vivo luminescence, 41 luminescence reaction, 37, 38 Bacterial luciferase, 33-35, 343 assay, 39 cloning, 34 crystal structure, 34 extraction and purification, 34 inactivation, 34, 35 molecular weight, 34 properties, 34 storage, 35 subunits, 34... 

W. Huang, A. Feltus, A. Witkowski, and S. Daunert, Homogeneous bioluminescence competitive binding assay for folate based on a coupled glucose-6-phosphate dehydrogenase-bacterial luciferase enzyme system. Anal. Chem. 68, 1646-1650 (1996). 

Chemical immobilization procedures of bioluminescent enzymes such as firefly luciferase and bacterial luciferase-NAD(P)H FMN oxidoreductase to glass beads or rods [174, 175], sepharose particles [176], and cellophane films [177] have produced active immobilized enzymes. Picomole-femtomole amounts of ATP or NAD(P)H could be detected using immobilized firefly luciferase or bacterial luciferase-oxidoreductase, respectively. 

The possibility of isolating the components of the two above-reported coupled reactions offered a new analytical way to determine NADH, FMN, aldehydes, or oxygen. Methods based on NAD(P)H determination have been available for some time and NAD(H)-, NADP(H)-, NAD(P)-dependent enzymes and their substrates were measured by using bioluminescent assays. The high redox potential of the couple NAD+/NADH tended to limit the applications of dehydrogenases in coupled assay, as equilibrium does not favor NADH formation. Moreover, the various reagents are not all perfectly stable in all conditions. Examples of the enzymes and substrates determined by using the bacterial luciferase and the NAD(P)H FMN oxidoreductase, also coupled to other enzymes, are listed in Table 5. 

B, bioluminescent bacterial system on nylon L, bioluminescent firefly system on nylon M, bioluminescent firefly system on methacrylate beads r-LM, recombinant firefly luciferase on methacrylate beads r-LN, recombinant firefly luciferase on nylon. 

Procedure 8.7 Bioluminescence assay of FMN using bacterial luciferase (EC... 

Methods based on chemiluminescent and bioluminescent labels are another area of nonisotopic immunoassays that continue to undergo active research. Most common approaches in this category are the competitive binding chemiluminescence immunoassays and the immunochemiluminometric assays. Chemiluminescence and heterogenous chemiluminescence immunoassays have been the subject of excellent reviews (91, 92). Detection in chemiluminescence immunoassays is based on either the direct monitoring of conjugated labels, such as luminol or acridinium ester, or the enzyme-mediated formation of luminescent products. Preparation of various derivatives of acridinium esters has been reported (93, 94), whereas a variety of enzyme labels including firefly or bacterial luciferase (70), horseradish peroxidase (86, 98), and alkaline phosphatase are commercially available. 

Production of light by certain marine bacteria. The general consensus is that light is produced when bacterial luciferase catalyzes the bioluminescent oxidation of FMNH2 and a long chain aldehyde by molecular oxygen. Volume 1(1,2). 

Most of bacterial biosensors are based on the operon luxCDABE that codes for the bacterial luciferase founded in the marine bacteria V. fischeri and V. harveyi, and for an essential aldehyde substrate that would otherwise have to be supplied exogenously. The cluster luxAB cassette codes for the luciferase whereas luxCDE encodes a fatty acid reductase complex. The latter enzymes are responsible for the synthesis of the long-chain aldehyde that is required as substrate in the bioluminescence reaction (Meighen and Dunlap, 1993 Hakkila et al., 2002). Luciferase catalyses the oxidation reaction of flavin mononucleotide (FMNH2). A long-chain (7 to 16 carbons) aldehyde is reduced in presence of oxygen by the aldehyde reductase. The outcome of the bioluminescent reaction can be expressed as follows ... 

Lee J. Sensitization by lumazine proteins of the bioluminescence emission ft om the reaction of bacterial luciferases. Photochem Photobiol 1982 36 689-97. 

Lin L, Szitmer R, Meighen E. Binding of flavin and aldehyde to the active site of bacterial luciferase. In Stanley P, Kricka L. eds. Bioluminescence Chemiluminescence Progress Current Applications. Singapore World Scientific, 2002 89-92. 

Figure 1. Bioluminescent emission spectra of bacterial luciferase from P. leiognathi in 1.48 % v/v acetone (1), 0.02 M pH 7.0 phosphate buffer (2) and 2.38 % v/v glycerol (3). Wavelengths of spectral maxima are indicated.

Bioluminescence-based analytical assays were used to measure various analytes in nanoliter sample volumes. Nanoliter volumes of multiple bioluminescent analytical assays were deposited in an array format and lyophilized. ATP-firefly luciferase (FFL) and NADH-bacterial luciferase (BL) platform reactions were compared. We achieved parallel sample delivery via sample-hydrated membranes. A CCD camera measured the luminescent kinetics for each assay. These miniaturized assays and instruments can he prepared as micro-analytical systems to operate in point-of-care (POC) diagnostic devices. 

Bacterial luciferase coimmobilized with NAD(P)H FMN oxidoreductase on starch gel has been used for bioluminescent assay of aldehydesCo-immobilization of bacterial luciferase, NAD(P)H FMN oxidoreductase and their substrates is referred to as multifunctional immobilized biosensor and is a new trend for use of bioluminescent analysis, e.g. toxicity biotest and bioassay. The main principle of this luciferase biotest is the correlation between toxicity of the sample being studied and changes in bioluminescence parameters in vitro. Toxicity of the sample is measured by the changes in bioliuninescence intensity compared with that of a control. Multifunctional immobilized biosensors based on luciferase have been used for the following bioassays. 

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