Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:1.14.14.3 (luciferase)
 38,195 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Luciferase from the anthozoan coelenterate Renilla reniformis (Renilla luciferin:oxygen 2-oxidoreductase (decarboxylating), EC 1.13.12.5.) catalyzes the bioluminescent oxidation of Renilla luciferin producing light (lambdaB 480 nm, QB 5.5%), oxyluciferin, and CO2 (Hori, K., Wampler, J.E., Matthews, J.C., and Cormier, M.J. (1973), Biochemistry 12, 4463). Using a combination of ion-exchange, molecular-sieve, sulfhydryl-exchange, and affinity chromatography, luciferase has been purified, approximately 12 000-fold with 24% recovery, to homogeneity as judged by analysis with disc and sodium dodecyl sulfate-polyacrylamide gel electrophoresis, gel filtration, and ultracentrifugation. Renilla luciferase is active as a nearly spherical single polypeptide chain monomer of 3.5 X 10(4) daltons having a specific activity of 1.8 X 10(15) hp s-1 mg-1 and a turnover number of 111 mumol min-1 mumol-1 of enzyme. This enzyme has a high content of aromatic and hydrophobic amino acids such that it has an epsilon280nm 0.1% of 2.1 and an average hydrophobicity of 1200 cal residue-1. The high average hydrophobicity of luciferase, which places it among the more hydrophobic proteins reported, is believed to account, at least in part, for its tendency to self-associate forming inactive dimers and higher molecular weight species.
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PMID:Purification and properties of Renilla reniformis luciferase. 1 97

A Ca2+-triggered luciferin-binding protein (BP-LH2) from the bioluminescent marine coelenterate, Renilla reniformis, has been purified by conventional methods. One kilogram of processed animals yields approximately 2.7 mg of pure protein with an overall yield of 55%. Physicochemical studies show that BP-LH2 is a globular protein containing one single polypeptide chain with one disulfide bond. Ultracentrifugation studies, amino acid analysis, and sodium dodecyl sulfate-gel electrophoresis show that BP-LH2 has an average molecular weight of 18,500. BP-LH2 has a Stokes radius of 23 A, a sedimentation coefficient, S020,w, of 2.3 S, and an isoelectric point of 4.3. The acidic nature of the protein was confirmed by amino acid analysis, which showed that 27% of the residues are acidic. The protein contains no carbohydrate, phosphate, or tryptophan. There is one noncovalently bound molecule of coelenterate type luciferin resulting in distinct protein spectral properties with absorption maxima at 276 nm (epsilon 0.1% 276 = 1.31) and 446 nm (episoln 0.1% 446 = 0.47) and a fluorescence emission at 520 nm (uncorrected). In the presence of Ca2+, BP-LH2 will react with Renilla luciferase to give the characteristic in vitro blue bioluminescence. Ca2+ binding produces a distinct change in the spectral properties of BP-LH2 including a 4-fold enhancement of tyrosine fluorescence at 332 nm and a 5-fold fluorescence enhancement at 520 nm. In addition, the visible absorption maximum shifts from 446 nm to 420 nm. The fluorescence enhancement at 320 nm occurs over the range from 1 to 10 micrometer Ca2+. BP-LH2 has two Ca2+-binding sites with an estimated Kd of 0.02 micrometer, in 10 muM Tris at pH 7.2. BP-LH2 was compared to several well studied Ca2+-binding proteins and was found to possess similar Ca2+-binding and physicochemical properties. This study clearly demonstrates that BP-LH2 is capable of triggering a bioluminescent flash in response to an intracellular Ca2+ transient.
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PMID:Ca2+-induced bioluminescence in Renilla reniformis. Purification and characterization of a calcium-triggered luciferin-binding protein. 3 74

Luciferase of the fireflies Luciola mingrelica was isolated from dried lanterns of fireflies and purified by chromatography on DEAE-Sephadex. The homogeneity of the preparation was determined by polyacrylamide gel disc electrophoresis. The molecular weight of the enzyme equal to 45000 was determined by disc electrophoresis in the presence of sodium dodecyl sulfate. The kinetic properties of the enzyme (V and Km for luciferin and ATP) within the pH-range of 7,0--8,5 were studied. The kinetic curves of the pH-dependences of log V and log Km for both substrates are bell-shaped, with a slope equal to 2. At pH optimum (7,7--7,9) the Km values for luciferin and ATP are 6,6 mkM and 0,3 mM, respectively. The properties of luciferase L. m. were compared to those of luciferase from fireflies Phophinus pyralis previously described in literature.
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PMID:[Kinetic properties of highly purified luciferase from fireflies Luciola mingrelica]. 4

The relationship between endogenous ATP and flagellar beat frequency of bull spermatozoa was investigated using cinematographic and biochemical criteria. ATP content was measured by the luciferin-luciferase method. Potassium, sodium, chloride, and viscosity were used to alter motility to establish the concomitant effects on ATP content. Beat frequency was correlated to endogenous ATP in potassium-supplemented media. Increased sodium concentration was related to ATP content, but not to beat frequency. With increasing viscosity of the medium, the frequency of flagellary beat decreased dramatically, while ATP content of the cells remained unchanged.
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PMID:Relationship of endogenous ATP to flagellar beat frequency in ejaculated bull spermatozoa. 15 23

A NAD(P)H:flavin oxidoreductase, which produces FMNH2, one of the substrates for the luciferase reaction in bioluminescent bacteria, has been purified with the aid of affinity chromatography on epsilon-aminohexanoyl-FMN-Sepharose. The purified enzyme, isolated from Beneckea harveyi, had a specific activity of 89 mumol of NADH oxidized/min/mg of protein at 23 degrees in the presence of saturating FMN and NADH and appeared homogeneous by several criteria on polyacrylamide gel electrophoresis. A molecular weight of 24,000 was estimated both by gel filtration and and sodium dodecyl sulfate gel electrophoresis indicating that the enzyme is composed of a single polypeptide chain. Kinetic studies showed that the higher specificity of the enzyme for NADH than NADPH and for riboflavin and FMN than FAD was primarily due to variations in the Michaelis constants for the different substrates. Initial velocity studies with all pairs of substrates gave intersecting patterns supporting a sequential mechanism for the NAD(P)H:flavin oxidoreductase.
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PMID:Purification and properties of a NAD(P)H:flavin oxidoreductase from the luminous bacterium, Beneckea harveyi. 30 40

The proteins of the bioluminescent bacterium Beneckea harveyi have been labelled with [3H]leucine prior to the induction of bioluminescence, and with [14C]leucine during the development of the bioluminescent system. An aliphatic aldehyde dehydrogenase and a NAD(P)H:flavin oxidoreductase, two enzymes that may be directly involved in the metabolism of the substrates (aldehyde, FMNH2) for the luminescent reaction catalyzed by luciferase, were purified and the isotope ratios of their respective polypeptide chains determined after sodium dodecyl sufate gel electrophoresis. A comparison of these isotope ratios to (a) the isotope ratios of the induced polypeptide chains of luciferase, purified in the same experiment, and (b) the average isotope ratio for the proteins synthesized in concert with growth has provided direct evidence that the synthesis of aldehyde dehydrogenase but not NAD(P)H:flavin oxidoreductase is induced during the development of bioluminescence.
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PMID:Differential synthesis of the polypeptides of aldehyde dehydrogenase and NAD(P)H:flavin oxidoreductase in the bioluminescent bacterium Beneckea harveyi. 30 25

The enzymatic activity of bacterial luciferase from Beneckea harveyi (a heterodimer, Mr = approximately 79,000) is rapidly lost upon treatment with trypsin or chymotrypsin. Under nondenaturing conditions, the proteolytically inactivated molecule has the same apparent molecular weight as the native enzyme, and appears to be relatively stable to further proteolytic degradation. Gel electrophoresis in sodium dodecyl sulfate of the products of this digestion shows that only the alpha subunit is degraded during the time of these experiments, and its rate of loss is the same as the rate of loss of light-producing activity. The action of either protease produces a species with mobility indicative of a molecular weight of about 28,000 and smaller fragments, and an unaltered beta subunit.
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PMID:Proteolytic inactivation of the luciferase from the luminous marine bacterium Beneckea harveyi. 30 51

Luminescent activity of spheroplasts of the cells of Photobacterium phosphoreum was stimulated by Rb+ and K+ and inhibited by Na+ in the medium. Opposite effects of these ions were observed on the rate of O2 consumption of the spheroplasts through the cytochrome and luciferase electron transfer systems. In vitro activities of NADH-FMN reductase and luciferase were only slightly stimulated by Rb+, K+, and Na+, while Na+ exhibited significant activation of the NADH-oxidizing activity of the cells. The redox level of cytochrome b in the spheroplasts during steady-state respiration in an Na+ medium was more reduced, while that of NAD(P) was more oxidized, than those in an Rb+ or K+ medium. Na+ activates the cytochrome electron transfer system at a point between NADH and cytochrome b, and thus has a stimulative effect on cellular O2 consumption. Rb+ and K+ do not show similar activation, but the cellular NAD(P) was brought to a more reduced level, so that in vivo luminescence is stimulated in an Rb+ or K+ medium.
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PMID:Luminescence and respiratory activities of Photobacterium phosphoreum. II. Control by monovalent cations. 59 51

The ligand specificity and biochemical properties of the human (h) FSH receptor are poorly characterized due to the low abundance of these receptors and the limited availability of human tissues. Using a fragment of rat FSH receptor cDNA, we screened a human testicular cDNA library and obtained a FSH receptor cDNA covering the entire amino acid-coding region. After transfection of a human fetal kidney cell line (293) with the hFSH receptor cDNA, radioligand receptor analysis revealed the presence of high affinity (Kd, 1.7 x 10(-9) M) FSH-binding sites on the plasma membrane. Both recombinant and wild-type hFSH displaced [125I]hFSH binding, with ED50 values of 25 and 70 ng/ml, respectively, whereas hLH, hCG, and hTSH were ineffective. Although human, rat(r), and ovine FSH as well as equine CG competed for rat testicular FSH receptor binding, only hFSH and rFSH interacted effectively with the recombinant hFSH receptor, suggesting that species-specific ligand recognition exists between human and rodent receptors. After incubation of transfected cells with hFSH, but not recombinant hLH or hCG, a dose-dependent increase (ED50, 10 ng/ml) in extracellular cAMP accumulation was observed, indicating a functional coupling of the expressed human receptor with the endogenous adenyl cyclase. In cells cotransfected with the FSH receptor expression plasmid and a luciferase reporter gene driven by the promoter of a cAMP-responsive gene, treatment with hFSH, but not hCG, resulted in a dose-dependent increase in luciferase activity. Northern blot analysis using a cRNA probe derived from the human receptor cDNA indicated the presence of multiple FSH receptor mRNA transcripts (7.0, 4.2, and 2.5 kilobases) in RNA prepared from human follicular phase ovary, but not from human corpus luteum or placenta. Additionally, two FSH-binding sites of 76 and 112 kilodaltons were detected in transfected 293 cells after ligand/receptor cross-linking and sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis. These results demonstrate the expression of functional hFSH receptor with unique ligand specificity and provide new data on the biochemical properties of the human receptor at the mRNA and protein levels.
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PMID:Expression of recombinant human follicle-stimulating hormone receptor: species-specific ligand binding, signal transduction, and identification of multiple ovarian messenger ribonucleic acid transcripts. 132 83

1. Stimulus-secretion coupling studies were carried out on adrenal chromaffin tissue from the toad. Catecholamine (CA) secretion was generated in response to acetylcholine (ACh) or high K+. 2. The response to ACh was found to be dependent on the presence of external Ca2+. The secretion induced by ACh or high K+ was inhibited by the Ca-channel blockers CoCl2 and nifedipine. 3. The specific Na+/H+ ionophore, monensin, induced a strong secretory response only if Na+ was present in the Ringer. Monensin's effect did not depend on external Ca2+ and was unaffected by the channel blockers tetrodotoxin or CoCl2. 4. Secretion induced by monensin was exocytotic as was shown by measuring ATP release using a photoluminescence, luciferine/luciferase assay. 5. In conclusion, in the toad, as in higher species, stimulus-secretion coupling involves Ca2+ entry from the external medium, possibly through voltage-dependent channels. Monensin is a potent secretagogue and the mechanism by which the ensuing elevation of intracellular Na+ concentration might induce a secretory response remains to be determined.
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PMID:Catecholamine secretion from adrenal chromaffin cells of the toad (Caudiverbera caudiverbera): effect of monensin. 135 94


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