60-92-4Relevant articles and documents
Structure and monomer/dimer equilibrium for the guanylyl cyclase domain of the optogenetics protein RhoGC
Kumar, Ramasamy P.,Morehouse, Benjamin R.,Fofana, Josiane,Trieu, Melissa M.,Zhou, Daniel H.,Lorenz, Molly O.,Oprian, Daniel D.
, p. 21578 - 21589 (2017)
RhoGC is a fusion protein from the aquatic fungus Blastocladiella emersonii, combining a type I rhodopsin domain with a guanylyl cyclase domain. It has generated excitement as an optogenetics tool for the manipulation of cyclic nucleotide signaling pathways. To investigate the regulation of the cyclase activity, we isolated the guanylyl cyclase domain from Escherichia coli with (GCwCCRho) and without (GCRho) the coiledcoil linker. Both constructs were constitutively active but were monomeric as determined by size-exclusion chromatography and analytical ultracentrifugation, whereas other class III nucleotidyl cyclases are functional dimers. We also observed that crystals of GCRho have only a monomer in an asymmetric unit. Dimers formed when crystals were grown in the presence of the non-cyclizable substrate analog 2′,3′-dideoxyguanosine- 5′-triphosphate, MnCl2, and tartrate, but their quaternary structure did not conform to the canonical pairing expected for class III enzymes. Moreover, the structure contained a disulfide bond formed with an active-site Cys residue required for activity. We consider it unlikely that the disulfide would form under intracellular reducing conditions, raising the possibility that this unusual dimer might have a biologically relevant role in the regulation of full-length RhoGC. Although we did not observe it with direct methods, a functional dimer was identified as the active state by following the dependence of activity on total enzymeconcentration. The low affinity observed for GCRhomonomers is unusual for this enzyme class and suggests that dimer formation may contribute to light activation of the full-length protein.
Photochemical Properties of New Photolabile cAMP Derivatives in a Physiological Saline Solution
Furuta, Toshiaki,Torigai, Hiromi,Sugimoto, Masazumi,Iwamura, Michiko
, p. 3953 - 3956 (1995)
Three new photolabile esters of cAMP (2-anthraquinonyl)methyl (1a), (7-methoxycoumarinyl)methyl (2a), and 2-naphthylmethyl (3a), have been developed.The stability and photochemical properties of these derivatives were compared to the previously reported ones in a physiological saline solution (1percent DMSO in Ringer's solution, pH 7.4).We found that 2a had satisfactory stability (t1/2 > 1000 h) in the dark and was photolyzed to release the parent cAMP on 340 nm irradiation (Φapp = 0.10, Ε340 = 6730) more efficiently than previously reported caged cAMPs.A biological test using the melanophores of the medaka (Oryzias latipes) revealed that 2a penetrated into the melanophores, inactive before irradiation and activated to release cAMP upon irradiation.We have developed a new caged cAMP which can be used in the investigation of biological responses regulated by intracellular cAMP concentrations using living cells.
ADENOSINE ANALOG AND ITS USE IN REGULATING THE CIRCADIAN CLOCK
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Paragraph 0166; 0171; 0172; 0208, (2018/08/12)
Provided are a kind of nucleoside analogue compounds, and compositions comprising these compounds and pentostatin, their use for modulating circadian rhythm, preferably, for shifting circadian phase, and methods for modulating circadian rhythm, preferably, for shifting circadian phase via these compounds or the compositions.
Factors influencing the operational stability of NADPH-dependent alcohol dehydrogenase and an NADH-dependent variant thereof in gas/solid reactors
Kulishova, Liliya,Dimoula, Kerasina,Jordan, Max,Wirtz, Astrid,Hofmann, Diana,Santiago-Schuebel, Beatrix,Fitter, Joerg,Pohl, Martina,Spiess, Antje C.
experimental part, p. 271 - 283 (2011/02/22)
The continuous enzymatic gas/solid bio-reactor serves both for the production of volatile fine chemicals and flavors on an industrial scale and for thermodynamically controlled investigation of substrate and water effects on enzyme preparations for research purposes. Here, we comparatively investigated the molecular effects on the operational stability of NADPH-dependent Lactobacillus brevis alcohol dehydrogenase and an NADH-dependent variant thereof, LbADH G37D, in the gas/solid bioreactor. The reference reaction is the reduction of acetophenone to (R)-1-phenylethanol with concomitant oxidation of 2-propanol to acetone for the purpose of regeneration of the redox cofactor. It could be clearly shown that not the thermostability of the cofactor, but the thermostability of the proteins in the solid dry state govern the order of magnitude of the operational stability of both purified enzymes in the gas/solid reactor at low thermodynamic activity of water and substrate. However, at higher thermodynamic activity the operational stability in the gas/solid reactor is overlaid by stabilizing and destabilizing effects of the substrates that require further investigation. We demonstrated first evidence that the substrate affinity of the two variants in the gas/solid reactor is similar to the affinity in aqueous medium. We could also show that partial unfolding of the proteins with subsequent aggregation are the factors governing protein thermo-in-stability both in the dissolved and in the dry state. Thus, stability investigations of enzymes in the dry state are suggested to predict their basal level of operational stability in gas/solid reactions.
