ABSTRACT
A model for the spatial structure of firefly luciferase--ATP--luciferin complex is suggested using the coordinates of unliganded luciferase and the enzyme--substrate complex of the adenylating subunit of gramicidin S synthetase known from the literature. Conformational changes in luciferase can occur during substrate binding resulting in a relative orientation of two luciferase domains similar to that in case of the AMP--phenylalanine--synthetase complex. The model is consistent with data on the physicochemical properties of firefly luciferase and its complexes with the substrates.
Subject(s)
Adenosine Triphosphate/metabolism , Luciferases/chemistry , Adenosine Triphosphate/chemistry , Animals , Binding Sites , Coleoptera/enzymology , Firefly Luciferin/metabolism , Luciferases/metabolism , Macromolecular Substances , Models, Molecular , Protein ConformationABSTRACT
Major advances in the development and application of the bioluminescent analysis to detect certain biologically active substances are discussed. The main merit of the method lies in its high sensitivity and specificity along with its simplicity and rapid performance. The available methodologies allow for detection of substances of varying nature: Ca2+, ATP, FMN, NAD(P), long-chain aldehydes, ATP- and NAD(P)-dependent enzymes and their substrates, many xenobiotics and antibiotics, and mutagens. The bioluminescence methodologies may be widely applied in clinical laboratory diagnosis.
Subject(s)
Luminescent Measurements , Adenosine Triphosphate/analysis , Aldehydes/analysis , Anti-Bacterial Agents/analysis , Firefly Luciferin/analysis , Flavin Mononucleotide/analysis , Humans , Luciferases/analysis , Models, Biological , Mutagens/analysis , NADP/analysis , Pancreatitis/enzymology , Protease Inhibitors/blood , Xenobiotics/analysisABSTRACT
Theoretical conformational analysis was carried out for a mixed disulfide of hemoglobin with glutathione. The conformational mobility of the beta-subunit C-terminal fragment in methemoglobin and deoxyhemoglobin either with free or glutathione-blocked reactive SH-groups was examined. The most stable conformations of the mixed disulfide were delineated. Its spatial structure was shown to be dependent on the hemoglobin state prior to the S-S bond formation: a disulfide made with deoxyhemoglobin had more closely interwoven hemoglobin and glutathione chains than the methemoglobin-derived disulfide. However, in both cases disulfide formation brought about the alterations in the spatial structure of hemoglobin as well as glutathione. The changes in the hemoglobin biochemical properties accompanying the association with glutathione were rationalized in the frames of the mixed disulfide structural analysis.
Subject(s)
Disulfides , Glutathione , Hemoglobins , Humans , Hydrogen Bonding , Models, Molecular , Molecular Conformation , Protein Conformation , Sulfhydryl CompoundsABSTRACT
Conformational analysis, by the method of atom-atomic potentials, has been carried out for five tripeptides containing gamma-glutamyl bonds and having general formula Glu(gamma)-X-Gly. The spatial structures have been determined and the changes arising on varying the second residue have been analyzed. A comparison of possible conformations and biological activity in respect to a number of enzymes allows to conceive what structural features of these compounds are important for the substrate specificity of the enzymes. In particular, the active site topography has been surmised for glutathione synthetase (EC 6.3.2.3) and gamma-glutamyltranspeptidase (EC 2.3.2.2). The glutathione thiol group has been found to be exposed in all possible conformations that explains its accessibility for various reagents.
