K. Turoverov - Academia.edu (original) (raw)
Papers by K. Turoverov
The Journal of Physical Chemistry A, 2007
The Journal of Physical Chemistry A, 2007
Journal of Applied Spectroscopy, 1978
Journal of Applied Spectroscopy, 2003
Comparative analysis of the absorption and fluorescence spectra and fluorescence excitation spect... more Comparative analysis of the absorption and fluorescence spectra and fluorescence excitation spectra of thioflavin T (ThT) in various solvents and in the composition of amyloid fibrils has shown that ThT, when excited in the region of the long-wavelength absorption band, fluoresces in the spectral region with a maximum at 478-484 nm. The appearance in aqueous and alcohol solutions of a fluorescence band with a maximum near 440 nm has been attributed to the presence in the composition of the ThT preparations of an impurity with an absorption band in the 340-350-nm range. The literature data showing that in glycerol ThT has a wide fluorescence spectrum with two maxima are due to the artifact connected with the use of a high concentration of the dye. It has been suggested that the cause of the low quantum yield of ThT aqueous and alcohol solutions is the breakage of the system of conjugated bonds due to the reorientation of the benzothiozole and benzaminic rings of ThT in the excited state with respect to one another. The main factor determining the high quantum yield of fluorescence of ThT incorporated in fibrils is the steric restriction of the rotation of the rings about one another under these conditions. The suggestions made have been verified by the quantumchemical calculation of the ThT molecule geometry in the ground and excited states.
Cell and Tissue Biology, 2007
A system of expression for the foreign actin gene in yeast cells Pichia pastoris has been develop... more A system of expression for the foreign actin gene in yeast cells Pichia pastoris has been developed. As a target protein, the Drosophila cytoplasmic actin 5C, which has 90% homology to the β -actin of higher eukaryotes, was used. In the present work, in order to develop conditions for biosynthesis of the target protein in yeast cells and a purification procedure for the recombinant protein, a GFP-actin fusion protein containing green fluorescent protein (GFP) as a fusion tag was expressed and purified. The size and survival of P. pastoris cells producing recombinant protein were characterized and shown to depend on the accumulation of recombinant actin. The purified fusion protein was used to obtain a polyclonal antibody necessary for testing for recombinant actin.
Biofizika
The localization of tryptophan residues in hen egg-white lysozyme macromolecule was studied on th... more The localization of tryptophan residues in hen egg-white lysozyme macromolecule was studied on the basis of the known 3D structure. The polarity and packing density of their microenvironments were evaluated. All residues that can affect the tryptophan fluorescence were revealed. It was shown that the orientation of these active groups relative to the indole ring of tryptophan plays a dramatic role in the efficiency of their influence. Tryptophan--tryptophan nonradiative energy transfer was evaluated from distances between tryptophan residues and their mutual orientation. The conformation of the side chains of tryptophan residues was determined. Special attention was paid to microenvironment of Trp108 responsible for the minor absorption band at 305 nm.
Journal of Applied Spectroscopy, 2014
ABSTRACT Spectral properties of a newly synthesized thiofl avin T (ThT) derivative, trans-2-[4-(d... more ABSTRACT Spectral properties of a newly synthesized thiofl avin T (ThT) derivative, trans-2-[4-(dimethylamino)styryl]-3-ethyl-1,3-benzothiazolium perchlorate (DMASEBT) with absorption and fluorescence spectra shifted to longer wavelengths (than ThT), were studied. Quantum-chemical calculations established that DMASEBT is planar in the ground state. The energy minimum of the excited molecule corresponded to a twisted conformation (TICT-state) with a 90° angle between the planar fragments. Charge in the molecule redistributed and a non-fluorescing TICT-state was formed if the fragments rotated. Fluorescence occurred from the non-equilibrium excited state (LE-state). It was shown that limited torsional rotation of the molecular fragments and; therefore, a decreased probability of transitioning into the non-fluorescing TICT-state, were responsible for the significantly increased quantum yield and fluorescence lifetime of DMASEBT upon increasing the solvent viscosity and incorporating it into amyloid fibrils.
Tsitologiia
The effect of filamentous (F) actin on the channel-forming activity of syringomycin E (SRE) in ne... more The effect of filamentous (F) actin on the channel-forming activity of syringomycin E (SRE) in negatively charged and uncharged bilayer lipid membranes (BLM) was studied. F-actin did not affect the membrane conductance in the absence of SRE. No changes in SRE-induced membrane conductance were observed when the above agents were added to the same side of BLM. However, the opposite side addition of F-actin and SRE provokes a multiple increase in membrane conductance. The similar voltage dependence of membrane conductance, equal values of single channel conductance and the effective gating charge of the channels upon F-actin action suggests that the actin-dependent increase in BLM conductance may result from an increase in the number of opened SRE-channels. BLM conductance kinetics depends on the sequence of SRE and F-actin addition, suggesting that actin-dependent rise of conductance may be induced by BLM structural changes that follow F-actin adsorption. F-actin exerted similar effec...
Journal of Applied Spectroscopy, 2015
ABSTRACT Absorption, fluorescence, and fluorescence excitation spectra of thioflavin T in aqueous... more ABSTRACT Absorption, fluorescence, and fluorescence excitation spectra of thioflavin T in aqueous solutions and polyvinyl alcohol films were studied. The dye aggregated when its concentration was increased. This was accompanied by a hypsochromic shift of the absorption spectrum and a bathochromic shift of the fluorescence spectrum. It was shown that fluorescence of the probe embedded in amyloid fibrils was caused by monomers rather than dimers or other aggregates.
![Research paper thumbnail of [Estimating of changes in the amyloid and prion content of yeast cells]](https://attachments.academia-assets.com/52556883/thumbnails/1.jpg)
](Tsitologiia, 2008
An attempt was made at estimating the overall amyloid content of yeast cells by treating crude ce... more An attempt was made at estimating the overall amyloid content of yeast cells by treating crude cellular lysates with thioflavin T, the agent specifically staining amyloid fibrils. We demonstrated that overproduction of the yeast chaperone Hsp104p, as well as GuHCI treatment of the [PSI+] cells led both to elimination of the [PSI+] factor and to a stable decrease of the overall amyloid content estimated by intensity of fluorescence (IF) of the thioflavin T. At the same time, overexpression of gene SUP35, coding the protein prionizable to [PSI+], led to generation of [PSI+] clones with higher IF of thioflavin T. Cytoduction in the crosses involving PSI factor leads to considerable enhancement of IF; cytoductants with the nucleus of the recipient [psi-] strain not only got [PSI+] factor from the donor strain but also increased their amyloid content. In these model experiments all treatments modifying one of the yeast prions, [PSI+] factor, led to a predictable shift of IF of thioflavin...
Tsitologiia, 2013
Investigation of the structure of ordered protein aggregates--amyloid fibrils, the influence of t... more Investigation of the structure of ordered protein aggregates--amyloid fibrils, the influence of the native structure of the protein and the environment on the process of fibrillation is currently the subject of intensive research. The present work is devoted to the study of the kinetics of insulin amyloid fibrils formation at low pH values (which are produced at many stages of the isolation and purification of the protein) using a fluorescent probe thioflavin T (ThT). It has been shown that the increase of fluorescence intensity of ThT during the formation of amyloid fibrils is described by a sigmoidal curve, in which 3 areas can be distinguished: the lag phase, the growth and the plateau, which characterize the various stages of fibril formation. Despite the variation in the length of the lag phase at the same experimental conditions (pH and temperature), we have found its reduction with stirring the solution and seeding. Data obtained using electron microscopy showed that the form...
Tsitologiia, 2013
The influence of various factors on the physico-chemical characteristics and complexation of gluc... more The influence of various factors on the physico-chemical characteristics and complexation of glucose with a mutant form of D-glucose/D-galactose-binding protein which can be regarded as a sensor of the glucometer, namely the protein GGBP/H152C with solvatochromic dye BADAN attached to the cysteine residue Cys 152, has been investigated. The point mutation His 152Cys and attaching BADAN reduced the affinity of the mutant form GGBP/H152C to glucose more than 8-fold compared to the wild type protein. This allows using this mutant for the determination of sugar content in biological fluids extracted by transdermal technologies. Sufficiently rapid complexation of GGBP/H152C with glucose (the time of protein-glucose complex formation is not more than three seconds even in solutions with a viscosity of 4 cP) provides timely monitoring changes in the concentration of sugar. The changes of ionic strength and pH within the physiological range of values of these variables do not have significa...
Tsitologiia, 2009
Recently it became evident that proteins can perform their function not only in globular state bu... more Recently it became evident that proteins can perform their function not only in globular state but also in partially or completely disordered state. The majority of globular proteins are enzymes which function is strictly determined. Regulation and signaling proteins participating in interconnection with variety of partners must have much more lability, and macromolecules of such proteins are mainly in partially or completely disordered state. The aim of this work was to describe from the unified viewpoint in the frame of energy landscape model the existence of native globular, native partially or completely disordered proteins, formation of intermolecular complexes with various partners, formation of amorphous aggregates and amyloid fibrils. Compact globular proteins are formed if polypeptide chain provides strong intramolecular interconnections. The ability of polypeptide chain to fold in a compact globule depends on the relation of hydrophobic and charged aminoacids in its compos...
Tsitologiia, 2007
Green fluorescent protein (GFP) from jellyfish Aequorea victoria is the most extensively studied ... more Green fluorescent protein (GFP) from jellyfish Aequorea victoria is the most extensively studied and widely used in cell biology protein. At present novel naturally occurring GFP-like proteins have been discovered and enhanced mutants of Aequorea GFP have been created. These mutants differ from wild-type GFP by stability, value of quantum yield, absorption and fluorescence spectra position and photochemical properties. GFP-like proteins are the fast growing family. This review is an attempt to characterize the main groups of GFP-like proteins, describe their structure and mechanisms of chromophore formation and summarize the main trends of their utilization as markers and biosensors in cell and molecular biology.
Tsitologiia, 2005
Results of actin folding-unfolding pathways examination and characterization of intermediate and ... more Results of actin folding-unfolding pathways examination and characterization of intermediate and misfolded states are summarized. Properties of microenvironments and peculiarities of location of tryptophan residues in protein are analysed in detail. This allowed to conclude that the main contribution to the bulk fluorescence of native protein is made by internal tryptophan residues Trp 340 and Trp 356, localized in hydrophobic regions, while tryptophan residues Trp 79 and Trp 86 are quenched. It has been shown that inactivated actin, previously regarded as an intermediate state between native and completely unfolded state of protein is in reality a misfolded aggregated state. The properties of actin in this state were characterized in detail. In particular, it is shown that inactivated actin is a monodisperse associate consisting of 15 monomer unit. Two earlier unknown intermediate states, which precede completely unfolding of protein macromolecule and formation of inactivated actin...
Tsitologiia, 2005
The present concepts of protein folding in vitro are reviewed. According to these concepts, amino... more The present concepts of protein folding in vitro are reviewed. According to these concepts, amino acid sequence of protein, which has appeared a result of evolutionary selection, determines the native structure of protein, the pathway of protein folding, and the existence of free energy barrier between native and denatured states of protein. The latter means that protein macromolecule can exist in either native or denatured state. And all macromolecules in the native state are identical but for structural fluctuations due to Brownian motion of their atoms. Identity of all molecules in native state is of primary importance for their correct functioning. The dependence of protein stability, which is measured as the difference between free energy of protein in native and denatured states, on temperature and denaturant concentration is discussed. The modern approaches characterizing transition state and nucleation are regarded. The role of intermediate and misfolded states in amorphous ...
Tsitologiia, 2005
The stability of fluorescent proteins (FPs) is of great importance for their use as reporters in ... more The stability of fluorescent proteins (FPs) is of great importance for their use as reporters in studies of gene expression, protein dynamics and localization in cell. A comparative analysis of conformational stability of fluorescent proteins, having different association state was done. The list of studied proteins includes EGFP (monomer of green fluorescent protein, GFP), zFP506 (tetramer GFP), mRFP1 and "dimer2" (monomer and dimmer of red fluorescent protein), DsRed1 (red tetramer). The character of fluorescence intensity changes induced by guanidine hydrochloride (GdnHCl) of these proteins differs significantly. Green tetramer zFP506 has been shown to be more stable than green monomer EGFP, red dimmer "dimer2" has been shown to be less stable than red tetramer DsRed1, while red monomer mRFP1 has been shown to be practically as stable as tetramer DsRedl. It is concluded that the quaternary structure, being an important stabilizing factor, does not represent th...
Tsitologiia, 2005
Unfolding--refolding of Escherichia coli disulfide isomerase C (DsbC) induced by GdnHCl was studi... more Unfolding--refolding of Escherichia coli disulfide isomerase C (DsbC) induced by GdnHCl was studied by intrinsic fluorescence. Interpretation of experimental fluorescence data was done together with the analysis of protein 3D structure. It is shown that although Cys 141 is the next neighbour of a single tryptophan residue Trp 140, sulfur atoms of the disulfide bond Cys 141--Cys 163 are far apart from the indole ring and cannot quench its fluorescence, while the potential quenchers are Met 136 and His 170. It has been revealed that, though each subunit of DsbC contains eight tyrosine residues, only three tyrosine residues (Tyr 171, Tyr 38 and Tyr 52) contribute to the bulk fluorescence of the molecule. The character of intrinsic fluorescence intensity changes induced by GdnHCl (equilibrium and kinetic data), the character of parametric dependencies between fluorescence intensity recorded at 320 and 365 nm, and the existence of an isosbestic point of protein fluorescence spectra in so...
The Journal of Physical Chemistry A, 2007
The Journal of Physical Chemistry A, 2007
Journal of Applied Spectroscopy, 1978
Journal of Applied Spectroscopy, 2003
Comparative analysis of the absorption and fluorescence spectra and fluorescence excitation spect... more Comparative analysis of the absorption and fluorescence spectra and fluorescence excitation spectra of thioflavin T (ThT) in various solvents and in the composition of amyloid fibrils has shown that ThT, when excited in the region of the long-wavelength absorption band, fluoresces in the spectral region with a maximum at 478-484 nm. The appearance in aqueous and alcohol solutions of a fluorescence band with a maximum near 440 nm has been attributed to the presence in the composition of the ThT preparations of an impurity with an absorption band in the 340-350-nm range. The literature data showing that in glycerol ThT has a wide fluorescence spectrum with two maxima are due to the artifact connected with the use of a high concentration of the dye. It has been suggested that the cause of the low quantum yield of ThT aqueous and alcohol solutions is the breakage of the system of conjugated bonds due to the reorientation of the benzothiozole and benzaminic rings of ThT in the excited state with respect to one another. The main factor determining the high quantum yield of fluorescence of ThT incorporated in fibrils is the steric restriction of the rotation of the rings about one another under these conditions. The suggestions made have been verified by the quantumchemical calculation of the ThT molecule geometry in the ground and excited states.
Cell and Tissue Biology, 2007
A system of expression for the foreign actin gene in yeast cells Pichia pastoris has been develop... more A system of expression for the foreign actin gene in yeast cells Pichia pastoris has been developed. As a target protein, the Drosophila cytoplasmic actin 5C, which has 90% homology to the β -actin of higher eukaryotes, was used. In the present work, in order to develop conditions for biosynthesis of the target protein in yeast cells and a purification procedure for the recombinant protein, a GFP-actin fusion protein containing green fluorescent protein (GFP) as a fusion tag was expressed and purified. The size and survival of P. pastoris cells producing recombinant protein were characterized and shown to depend on the accumulation of recombinant actin. The purified fusion protein was used to obtain a polyclonal antibody necessary for testing for recombinant actin.
Biofizika
The localization of tryptophan residues in hen egg-white lysozyme macromolecule was studied on th... more The localization of tryptophan residues in hen egg-white lysozyme macromolecule was studied on the basis of the known 3D structure. The polarity and packing density of their microenvironments were evaluated. All residues that can affect the tryptophan fluorescence were revealed. It was shown that the orientation of these active groups relative to the indole ring of tryptophan plays a dramatic role in the efficiency of their influence. Tryptophan--tryptophan nonradiative energy transfer was evaluated from distances between tryptophan residues and their mutual orientation. The conformation of the side chains of tryptophan residues was determined. Special attention was paid to microenvironment of Trp108 responsible for the minor absorption band at 305 nm.
Journal of Applied Spectroscopy, 2014
ABSTRACT Spectral properties of a newly synthesized thiofl avin T (ThT) derivative, trans-2-[4-(d... more ABSTRACT Spectral properties of a newly synthesized thiofl avin T (ThT) derivative, trans-2-[4-(dimethylamino)styryl]-3-ethyl-1,3-benzothiazolium perchlorate (DMASEBT) with absorption and fluorescence spectra shifted to longer wavelengths (than ThT), were studied. Quantum-chemical calculations established that DMASEBT is planar in the ground state. The energy minimum of the excited molecule corresponded to a twisted conformation (TICT-state) with a 90° angle between the planar fragments. Charge in the molecule redistributed and a non-fluorescing TICT-state was formed if the fragments rotated. Fluorescence occurred from the non-equilibrium excited state (LE-state). It was shown that limited torsional rotation of the molecular fragments and; therefore, a decreased probability of transitioning into the non-fluorescing TICT-state, were responsible for the significantly increased quantum yield and fluorescence lifetime of DMASEBT upon increasing the solvent viscosity and incorporating it into amyloid fibrils.
Tsitologiia
The effect of filamentous (F) actin on the channel-forming activity of syringomycin E (SRE) in ne... more The effect of filamentous (F) actin on the channel-forming activity of syringomycin E (SRE) in negatively charged and uncharged bilayer lipid membranes (BLM) was studied. F-actin did not affect the membrane conductance in the absence of SRE. No changes in SRE-induced membrane conductance were observed when the above agents were added to the same side of BLM. However, the opposite side addition of F-actin and SRE provokes a multiple increase in membrane conductance. The similar voltage dependence of membrane conductance, equal values of single channel conductance and the effective gating charge of the channels upon F-actin action suggests that the actin-dependent increase in BLM conductance may result from an increase in the number of opened SRE-channels. BLM conductance kinetics depends on the sequence of SRE and F-actin addition, suggesting that actin-dependent rise of conductance may be induced by BLM structural changes that follow F-actin adsorption. F-actin exerted similar effec...
Journal of Applied Spectroscopy, 2015
ABSTRACT Absorption, fluorescence, and fluorescence excitation spectra of thioflavin T in aqueous... more ABSTRACT Absorption, fluorescence, and fluorescence excitation spectra of thioflavin T in aqueous solutions and polyvinyl alcohol films were studied. The dye aggregated when its concentration was increased. This was accompanied by a hypsochromic shift of the absorption spectrum and a bathochromic shift of the fluorescence spectrum. It was shown that fluorescence of the probe embedded in amyloid fibrils was caused by monomers rather than dimers or other aggregates.
Tsitologiia, 2008
An attempt was made at estimating the overall amyloid content of yeast cells by treating crude ce... more An attempt was made at estimating the overall amyloid content of yeast cells by treating crude cellular lysates with thioflavin T, the agent specifically staining amyloid fibrils. We demonstrated that overproduction of the yeast chaperone Hsp104p, as well as GuHCI treatment of the [PSI+] cells led both to elimination of the [PSI+] factor and to a stable decrease of the overall amyloid content estimated by intensity of fluorescence (IF) of the thioflavin T. At the same time, overexpression of gene SUP35, coding the protein prionizable to [PSI+], led to generation of [PSI+] clones with higher IF of thioflavin T. Cytoduction in the crosses involving PSI factor leads to considerable enhancement of IF; cytoductants with the nucleus of the recipient [psi-] strain not only got [PSI+] factor from the donor strain but also increased their amyloid content. In these model experiments all treatments modifying one of the yeast prions, [PSI+] factor, led to a predictable shift of IF of thioflavin...
Tsitologiia, 2013
Investigation of the structure of ordered protein aggregates--amyloid fibrils, the influence of t... more Investigation of the structure of ordered protein aggregates--amyloid fibrils, the influence of the native structure of the protein and the environment on the process of fibrillation is currently the subject of intensive research. The present work is devoted to the study of the kinetics of insulin amyloid fibrils formation at low pH values (which are produced at many stages of the isolation and purification of the protein) using a fluorescent probe thioflavin T (ThT). It has been shown that the increase of fluorescence intensity of ThT during the formation of amyloid fibrils is described by a sigmoidal curve, in which 3 areas can be distinguished: the lag phase, the growth and the plateau, which characterize the various stages of fibril formation. Despite the variation in the length of the lag phase at the same experimental conditions (pH and temperature), we have found its reduction with stirring the solution and seeding. Data obtained using electron microscopy showed that the form...
Tsitologiia, 2013
The influence of various factors on the physico-chemical characteristics and complexation of gluc... more The influence of various factors on the physico-chemical characteristics and complexation of glucose with a mutant form of D-glucose/D-galactose-binding protein which can be regarded as a sensor of the glucometer, namely the protein GGBP/H152C with solvatochromic dye BADAN attached to the cysteine residue Cys 152, has been investigated. The point mutation His 152Cys and attaching BADAN reduced the affinity of the mutant form GGBP/H152C to glucose more than 8-fold compared to the wild type protein. This allows using this mutant for the determination of sugar content in biological fluids extracted by transdermal technologies. Sufficiently rapid complexation of GGBP/H152C with glucose (the time of protein-glucose complex formation is not more than three seconds even in solutions with a viscosity of 4 cP) provides timely monitoring changes in the concentration of sugar. The changes of ionic strength and pH within the physiological range of values of these variables do not have significa...
Tsitologiia, 2009
Recently it became evident that proteins can perform their function not only in globular state bu... more Recently it became evident that proteins can perform their function not only in globular state but also in partially or completely disordered state. The majority of globular proteins are enzymes which function is strictly determined. Regulation and signaling proteins participating in interconnection with variety of partners must have much more lability, and macromolecules of such proteins are mainly in partially or completely disordered state. The aim of this work was to describe from the unified viewpoint in the frame of energy landscape model the existence of native globular, native partially or completely disordered proteins, formation of intermolecular complexes with various partners, formation of amorphous aggregates and amyloid fibrils. Compact globular proteins are formed if polypeptide chain provides strong intramolecular interconnections. The ability of polypeptide chain to fold in a compact globule depends on the relation of hydrophobic and charged aminoacids in its compos...
Tsitologiia, 2007
Green fluorescent protein (GFP) from jellyfish Aequorea victoria is the most extensively studied ... more Green fluorescent protein (GFP) from jellyfish Aequorea victoria is the most extensively studied and widely used in cell biology protein. At present novel naturally occurring GFP-like proteins have been discovered and enhanced mutants of Aequorea GFP have been created. These mutants differ from wild-type GFP by stability, value of quantum yield, absorption and fluorescence spectra position and photochemical properties. GFP-like proteins are the fast growing family. This review is an attempt to characterize the main groups of GFP-like proteins, describe their structure and mechanisms of chromophore formation and summarize the main trends of their utilization as markers and biosensors in cell and molecular biology.
Tsitologiia, 2005
Results of actin folding-unfolding pathways examination and characterization of intermediate and ... more Results of actin folding-unfolding pathways examination and characterization of intermediate and misfolded states are summarized. Properties of microenvironments and peculiarities of location of tryptophan residues in protein are analysed in detail. This allowed to conclude that the main contribution to the bulk fluorescence of native protein is made by internal tryptophan residues Trp 340 and Trp 356, localized in hydrophobic regions, while tryptophan residues Trp 79 and Trp 86 are quenched. It has been shown that inactivated actin, previously regarded as an intermediate state between native and completely unfolded state of protein is in reality a misfolded aggregated state. The properties of actin in this state were characterized in detail. In particular, it is shown that inactivated actin is a monodisperse associate consisting of 15 monomer unit. Two earlier unknown intermediate states, which precede completely unfolding of protein macromolecule and formation of inactivated actin...
Tsitologiia, 2005
The present concepts of protein folding in vitro are reviewed. According to these concepts, amino... more The present concepts of protein folding in vitro are reviewed. According to these concepts, amino acid sequence of protein, which has appeared a result of evolutionary selection, determines the native structure of protein, the pathway of protein folding, and the existence of free energy barrier between native and denatured states of protein. The latter means that protein macromolecule can exist in either native or denatured state. And all macromolecules in the native state are identical but for structural fluctuations due to Brownian motion of their atoms. Identity of all molecules in native state is of primary importance for their correct functioning. The dependence of protein stability, which is measured as the difference between free energy of protein in native and denatured states, on temperature and denaturant concentration is discussed. The modern approaches characterizing transition state and nucleation are regarded. The role of intermediate and misfolded states in amorphous ...
Tsitologiia, 2005
The stability of fluorescent proteins (FPs) is of great importance for their use as reporters in ... more The stability of fluorescent proteins (FPs) is of great importance for their use as reporters in studies of gene expression, protein dynamics and localization in cell. A comparative analysis of conformational stability of fluorescent proteins, having different association state was done. The list of studied proteins includes EGFP (monomer of green fluorescent protein, GFP), zFP506 (tetramer GFP), mRFP1 and "dimer2" (monomer and dimmer of red fluorescent protein), DsRed1 (red tetramer). The character of fluorescence intensity changes induced by guanidine hydrochloride (GdnHCl) of these proteins differs significantly. Green tetramer zFP506 has been shown to be more stable than green monomer EGFP, red dimmer "dimer2" has been shown to be less stable than red tetramer DsRed1, while red monomer mRFP1 has been shown to be practically as stable as tetramer DsRedl. It is concluded that the quaternary structure, being an important stabilizing factor, does not represent th...
Tsitologiia, 2005
Unfolding--refolding of Escherichia coli disulfide isomerase C (DsbC) induced by GdnHCl was studi... more Unfolding--refolding of Escherichia coli disulfide isomerase C (DsbC) induced by GdnHCl was studied by intrinsic fluorescence. Interpretation of experimental fluorescence data was done together with the analysis of protein 3D structure. It is shown that although Cys 141 is the next neighbour of a single tryptophan residue Trp 140, sulfur atoms of the disulfide bond Cys 141--Cys 163 are far apart from the indole ring and cannot quench its fluorescence, while the potential quenchers are Met 136 and His 170. It has been revealed that, though each subunit of DsbC contains eight tyrosine residues, only three tyrosine residues (Tyr 171, Tyr 38 and Tyr 52) contribute to the bulk fluorescence of the molecule. The character of intrinsic fluorescence intensity changes induced by GdnHCl (equilibrium and kinetic data), the character of parametric dependencies between fluorescence intensity recorded at 320 and 365 nm, and the existence of an isosbestic point of protein fluorescence spectra in so...