Activity and fluorescence changes of lactate dehydrogenase induced by guanidine hydrochloride in reverse micelles

Abstract

Denaturants activate several multimeric enzymes in reverse micelles [Garza-Ramos, G., Darszon, A., Tuena de G6mez-Puyou, M. & G6mez-Puyou, A. (1992) Eul: J. Biochem. 205, 509-5171. Here, the effect on activity and intrinsic fluorescence of pig heart lactate dehydrogenase (LDH) in reverse micelles [formed with 0.2 M cetyltrimethylammonium bromide in octanehexanol (8.6: 1, by vol.)] was explored at various water and guanidine hydrochloride (GdnMC1) concentrations. Emission fluorescence spectra of LDH in aqueous media and in micelles were similar. As in all aqueous media, 1.0 M Gdn/HCl in the water phase of reverse micelles produced fluorescence quenching and a blue shift of the maximal emission. In 5.0 M Gdn/HCl, instead of the red shift and significant quenching seen in water, the maximum emission further shifted to the blue and was only slightly quenched. Gdn/HCl titrations of activity and fluorescence changes of LDH in micelles with different water contents showed that at Wo

Spectroelectrochemical Investigation of 4-Dicyanomethylene 2,6-Dimethyl-4H-Pyran (DDP) Dye with Guanidine Hydrochloride (GuHCl) in Water

“…GuHCl forms hydrogen-bonding with water molecule through N-H hydrogen and acts as a hydrogen-bonding donor only wherein urea acts as a hydrogen-bonding donor as well as an acceptor (Kawahara & Tanford, 1966). It has been established that GuHCl is surrounded approximately by 12 to 13 water molecules and six water molecules are involved in a direct hydrogen-bonding interaction with GuHCl (Castellino & Barker, 1968;Garza-Ramos et al, 1992;Strambini & Gonnelli, 1986;Ma & Tsou, 1991;Miggiano, Mordente, Pischiutta, Martorana, & Castelli, 1987;Zettl meissl, Rudolph, & Jaenicke, 1982;Moosavi-Movahedi, Naderi, & Farzami, 1994;Morjana, McKeone, & Gilbert, 1993;Shoshani, Darszon, Tijena de Gomez-puyou, & Gomez-puyou, 1994;Cox, 1968;Katz, 1968;Makhatadze & Privalov, 1992;Hedwig, Lilley, & Linsdell, 1991;Michnik & Sulkowska, 1997). This variation in the hydrogen-bonding pattern compared to urea is significantly important because the nature and the type of hydrogen-bonding arrangement influence the photophysical properties of fluorescence probes.…”

“…An increase in the concentration of GuHCl results in the partial displacement of the water molecules from the secondary hydration sphere resulting in a GuHCl-water hydrogen-bonding network throughout the medium. This results in a large variation in the orientation of the hydrophobic and hydrophilic clusters in aqueous phase in the close vicinity of GuHCl (Castellino & Barker, 1968;Moosavi-Movahedi, Naderi, & Farzami, 1994;Morjana, McKeone, & Gilbert, 1993;Shoshani, Darszon, Tijena de Gomez-puyou, & Gomez-puyou, 1994;Cox, 1968).…”

“…Apart from the solvent mediated hydrogen-bonding interactions, the electrostatic interactions in combination with hydrogen-bonding influence the excited state properties of probes. GuHCl and urea both possess strong hydrogen-bonding properties with solvents and have a tendency to alter the structure of the water molecules which is crucial in biological studies concerned to GuHCl (Strambini & Gonnelli, 1986;Zettl meissl, Rudolph, & Jaenicke, 1982;Moosavi-Movahedi, Naderi, & Farzami, 1994;Morjana, McKeone, & Gilbert, 1993;Shoshani, Darszon, Tijena de Gomez-puyou, & Gomez-puyou, 1994;Cox, 1968;Katz, 1968;Makhatadze & Privalov, 1992;Hedwig, Lilley, & Linsdell, 1991). The variation in the photophysical properties of probes with strong chaotropic agents is an area of unexplored domain and the literature regarding the photophysical properties of an extrinsic fluorescent probe with GuHCl is reported less in literature compared to that of urea interaction with fluorescent probes in aqueous solution (Kumaran, Varalakshmi, & Padma Malar, 2010;Sowntharya, Gayathri, Dhenadayalan, Vasanthi, Vanjinathan, & Kumaran, 2017;Kawahara & Tanford, 1966;Castellino & Barker, 1968;Garza-Ramos et al, 1992;Strambini & Gonnelli, 1986).…”

Effect of urea on the enzymatic activity of a lipase entrapped in AOT–heptane–water reverse micellar solutions

“…To the 0021-9797/$ -see front matter  2004 Elsevier Inc. All rights reserved. doi:10.1016/j.jcis.2004.08.176 best of our knowledge, there are very few reported studies addressing the issue of the effect of denaturants on the activities of enzymes in reverse micellar solutions [17][18][19]. Such studies are important since the denaturant in the micellar solution could lead either to a resistance to denaturation, if the enzyme is stabilized by the micellar interface, or to an enhanced effect, relative to the aqueous solution, if the enzyme is subjected to enlarged inactivation due to the presence of urea at the micellar interface.…”