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Biomedical subjects

L I Krishtalik

Publications and source records attributed to L I Krishtalik.

At least 19 recordsLinked to original sources
PubMed2001

Photosynthetic electron transfer controlled by protein relaxation: analysis by Langevin stochastic approach.

Relaxation processes in proteins range in time from picoseconds to seconds. Correspondingly, biological electron transfer (ET) could be controlled by slow protein relaxation. We used the Langevin stochastic approach to describe this type of ET dynamics. Two different types of kinetic behavior were revealed, namely: oscillating ET (that could occur at picoseconds) and monotonically relaxing ET. On a longer time scale, the ET dynamics can include two different kinetic components. The faster one reflects the initial, nonadiabatic ET, whereas the slower one is governed by the medium relaxation. We derived a simple relation between the relative extents of these components, the change in the free energy (DeltaG), and the energy of the slow reorganization Lambda. The rate of ET was found to be determined by slow relaxation at -DeltaG < or = Lambda. The application of the developed approach to experimental data on ET in the bacterial photosynthetic reaction centers allowed a quantitative description of the oscillating features in the primary charge separation and yielded values of Lambda for the slower low-exothermic ET reactions. In all cases but one, the obtained estimates of Lambda varied in the range of 70-100 meV. Because the vast majority of the biological ET reactions are only slightly exothermic (DeltaG > or = -100 meV), the relaxationally controlled ET is likely to prevail in proteins.

Crystallography, X-Ray↗
PubMed2000

Effects of medium polarization and pre-existing field on activation energy of enzymatic charge-transfer reactions.

The highly organized spatial structure of proteins' polar groups results in the existence of a permanent intraprotein electric field and in protein's weak dielectric response, i.e. its low dielectric constant. The first factor affects equilibrium free energy gap of a charge-transfer reaction, the second (medium polarization effect) influences both equilibrium and non-equilibrium (reorganization) energies, decreasing the latter substantially. In the framework of the rigorous 'fixed-charge-density' formalism, the medium polarization component of the reaction activation energy has been calculated, both for the activation energy of the elementary act proper, and the effective activation energy accounting for the charges' transfer from water into a low-dielectric structureless medium. In all typical cases of reactions, the energy spent for charge transfer from water into structureless 'protein' is larger than the gain in activation energy due to the protein's low reorganization energy. Therefore, the low dielectric constant of proteins is not sufficient to ensure their high catalytic activity, and an additional effect of the pre-existing intraprotein electric field, compensating for an excessive charging energy, is necessary. Only a combined action of low reorganization energy and pre-existing electric field provides proteins with their high catalytic activity. The dependence of activation energy on the globule geometry has been analyzed. It is shown that, for each reaction, an optimum set of geometric parameters exists. For five hydrolytic enzymes, the optimum globule radii have been calculated using the experimental geometry of their active sites. The calculated radii agree satisfactorily with the real sizes of these macromolecules, both by absolute and by relative values.

Carboxypeptidases↗
PubMed2000

The mechanism of the proton transfer: an outline.

A brief summary of the principal notions of the quantum-mechanical theory of the charge transfer reactions has been presented. In the framework of this theory, the mechanism of the proton transfer consists in the classical medium reorganization that equalizes the proton energy levels in the initial and final states, and a consequent proton transfer via a quantum-mechanical underbarrier transition. On the basis of this mechanism, factors influencing the proton transfer probability, and hence kinetic isotope effect, have been discussed; among them are the optimum tunneling distance, the involvement of the excited vibrational states, etc. Semi-classical and quantum-mechanical treatments of the Swain-Schaad relations have been compared. Some applications to enzymatic proton-transfer reactions have been described.

Energy Metabolism↗
PubMed2000

Low dielectric response in enzyme active site.

The kinetics of charge transfer depend crucially on the dielectric reorganization of the medium. In enzymatic reactions that involve charge transfer, atomic dielectric response of the active site and of its surroundings determines the efficiency of the protein as a catalyst. We report direct spectroscopic measurements of the reorganization energy associated with the dielectric response in the active site of alpha-chymotrypsin. A chromophoric inhibitor of the enzyme is used as a spectroscopic probe. We find that water strongly affects the dielectric reorganization in the active site of the enzyme in solution. The reorganization energy of the protein matrix in the vicinity of the active site is similar to that of low-polarity solvents. Surprisingly, water exhibits an anomalously high dielectric response that cannot be described in terms of the dielectric continuum theory. As a result, sequestering the active site from the aqueous environment inside low-dielectric enzyme body dramatically reduces the dielectric reorganization. This reduction is particularly important for controlling the rate of enzymatic reactions.

Binding Sites↗
PubMed1999

Free energy of charge transfer and intraprotein electric field: method of calculation depends on the charge state of protein at a given structure.

Free energy of charge transfer presents a basic characteristic of reactions such as protonation, oxido-reduction and similar. Evaluation of this quantity requires calculation of charging energy. Proteins are structured dielectrics, and a consistent incorporation of their structure into calculation of intraprotein electric field results in expression for charging energy of an active group in protein, which is essentially different from that for a simple dielectric. An algorithm for semi-continuum calculation of relevant free energies is described. First of the two components of charging energy in protein, energy of the medium response to charge redistribution in reactants, should be always calculated as the charging energy by the charge redistribution using the static dielectric constant of protein. The second term is interaction energy of the charge redistribution with the 'frozen' electric field of the system before reaction. Charges of protein groups, at which the protein structure has been determined, are often different from those before reaction of charge transfer, so is the corresponding intraprotein field. The field is expressed through either both the optical and static dielectric constants of protein or only optical one depending on whether the charges of protein groups before reaction and upon structural analysis are the same or not. Proper allowance for difference in charges of reacting groups before reaction and upon structural analysis of protein is thermodynamically necessary and quantitatively important. The expression for activation free energy for charge transfer in proteins is derived in the form presenting explicitly an invariant contribution of protein structure.

Algorithms↗
PubMed1997

Electrostatic effects on electron-transfer kinetics in the cytochrome f-plastocyanin complex.

In a complex of two electron-transfer proteins, their redox potentials can be shifted due to changes in the dielectric surroundings and the electrostatic potentials at each center caused by the charged residues of the partner. These effects are dependent on the geometry of the complex. Three different docking configurations (DCs) for intracomplex electron transfer between cytochrome f and plastocyanin were studied, defined by 1) close contact of the positively charged region of cytochrome f and the negatively charged regions of plastocyanin (DC1) and by (2, 3) close contact of the surface regions adjacent to the Fe and Cu redox centers (DC2 and DC3). The equilibrium energetics for electron transfer in DC1-DC3 are the same within approximately +/-0.1 kT. The lower reorganization energy for DC2 results in a slightly lower activation energy for this complex compared with DC1 and DC3. The long heme-copper distance (approximately 24 A) in the DC1 complex drastically decreases electronic coupling and makes this complex much less favorable for electron transfer than DC2 or DC3. DC1-like complexes can only serve as docking intermediates in the pathway toward formation of an electron-transfer-competent complex. Elimination of the four positive charges arising from the lysine residues in the positive patch of cytochrome f, as accomplished by mutagenesis, exerts a negligible effect (approximately 3 mV) on the redox potential difference between cyt f and PC.

Biophysical Phenomena↗
PubMed1997

Electrostatics of proteins: description in terms of two dielectric constants simultaneously.

In the semi-continuum treatment of the energetics of charge formation (or transfer) inside a protein, two components of the energy are inevitably present: the energy of interaction of the ion with the pre-existing intraprotein electric field, and the energy due to polarization of the medium by the newly formed charge. The pre-existing field is set up by charges (partial or full) of the protein atoms fixed in a definite structure. The calculation of this field involves only the electronic polarization (the optical dielectric constant epsilon(o)) of the protein because the polarization due to shifts of heavy atoms has already been accounted for by their equilibrium coordinates. At the same time, the aqueous surroundings should be described by the static constant epsilon(sw), as the positions of water molecules are not fixed. The formation of a new charge, absent in the equilibrium X-ray structure, results in shifts of electrons and polar atoms, i.e., it involves all kinds of medium polarization described by the static dielectric constant of protein epsilon(s). Thus, in calculations of the total energy, two different dielectric constants of the protein are operative simultaneously. This differs from a widely used algorithm employing one effective dielectric constant for both components of the ion's energy.

Proteins↗
PubMed1995

On the physical basis for the cis-positive rule describing protein orientation in biological membranes.

The topology of hydrophobic intramembrane proteins is characterized by a statistical asymmetry in the distribution of positively-charged residues on the two sides of the membrane, the 'inside- or cis-positive rule'. A mechanism is proposed involving only neutral residue transfer. For a tightly bound polypeptide adsorbed on the membrane and not at equilibrium, the pK values of the ionic residues related to dissociation of the proton into the aqueous phase bulk are increased because of interaction with the negative charges at the membrane surface. The pK shift would selectively neutralize aspartate and glutamate residues, favoring their translocation across the membrane, while stabilizing the impermeant positively charged state of lysine and arginine residues.

Cell Membrane↗
PubMed1993

The redox properties of cytochromes b imposed by the membrane electrostatic environment.

The effect of the dipole potential field of extended membrane spanning alpha-helices on the redox potentials of b cytochromes in energy transducing membranes has been calculated in the context of a three phase model for the membrane. In this model, the membrane contains three dielectric layers; (i) a 40-A hydrophobic membrane bilayer, with dielectric constant em = 3-4, (ii) 10-20-A interfacial layers of intermediate polarity, ein = 12-20, that consist of lipid polar head groups and peripheral protein segments, and (iii) an external infinite water medium, ew = 80. The unusually positive midpoint potential, Em = +0.4 V, of the "high potential" cytochrome b-559 of oxygenic photosynthetic membranes, a previously enigmatic property of this cytochrome, can be explained by (i) the position of the heme in the positive dipole potential region near the NH2 termini of the two parallel helices that provide its histidine ligands, and (ii) the loss of solvation energy of the heme ion due to the low dielectric constant of its surroundings, leading to an estimate of +0.31 to +0.37 V for the cytochrome Em. The known tendency of this cytochrome to undergo a large -delta Em shift upon exposure of thylakoid membranes to proteases or damaging treatments is explained by disruption of the intermediate polarity (ein) surface dielectric layer and the resulting contact of the heme with the external water medium. Application of this model to the two hemes (bn and bp) of cytochrome b of the cytochrome bc1 complex, with the two hemes placed symmetrically in the low dielectric (em) membrane bilayer, results in Em values of hemes bn and bp that are, respectively, somewhat too negative (approximately -0.1 V), and much too positive (approximately +0.3 V), leading to a potential difference, Em(bp) - Em(bn), with the wrong sign and magnitude, +0.25 V instead of -0.10 to -0.15 V. The heme potentials can only be approximately reconciled with experiment, if it is assumed that the two hemes are in different dielectric environments, with that of heme bp being more polar.

Amino Acid Sequence↗
PubMed1985

Conformational mobility of proteins and activation energy of enzymatic charge transfer reactions: an analysis in dielectric formalism.

In calculating the medium reorganization energy and the activation energy of charge transfer enzymatic reactions, an allowance is made for the enhanced conformational mobility of the protein external region. The two-layer model is proposed, the outer layer having a higher static dielectric constant. The calculations show that the higher mobility in the outer layer causes some quantitative rather than qualitative changes. The main result obtained earlier is confirmed: the reorganization energy for charge transfer reaction in protein globule is much lower than in water and for this reason the activation energy also decreases. The higher dielectric constant of the outer layer somewhat favours the introduction of charge into active site and hence favours the natural selection of proteins as enzymes. This effect cannot exclude the necessity of other factors stabilizing ionic forms inside the protein globule. Freezing of conformational mobility (say, at low temperatures) hinders the charge transfer process as a consequence of the difficulty in equalizing the initial and final energy levels.

Binding Sites↗
PubMed1985

Medium reorganization energy and enzymatic reaction activation energy.

Reorganization and activation energies for charge transfer reactions occurring inside a dielectric sphere have been calculated by solving the problem of polar medium reorganization within and outside a dielectric sphere placed in another infinite dielectric. The dielectric sphere is assumed to simulate a protein globule, i.e. an enzyme molecule. It has been shown that for some reaction types the activation energy tends to decrease as the globule radius increases and that for each of the reaction types considered there is an optimal globule radius an increase of which does not bring about any tangible activation energy reduction. The calculated optimal radii for different processes are in good agreement with the increasing molecular sizes in the series: ribonuclease less than or equal to lysozyme less than serine proteinases approximately equal to cysteine proteinases less than NAD-dependent dehydrogenases. The calculated radii are usually about 1.5 to 1.7 times (and molecular masses about 4-5 times) smaller than the experimental ones. The reasons for this discrepancy are discussed and it has been suggested that the approximate nature of the treatment of a protein globule as a structureless dielectric is the main reason. It is shown that charge transfer at an acute angle to the globule surface is the optimum process. For endoergonic reaction stages it is the net charge transfer towards the periphery and for exoergonic ones that in the reverse direction which are advantageous. These conclusions are consistent with the data about the structure of the above-mentioned enzymes.

Alcohol Dehydrogenase↗
PubMed1985

Effective activation energy of enzymatic and nonenzymatic reactions. Evolution-imposed requirements to enzyme structure.

The difference of the activation energies in a protein globule and water has been treated in terms of the theory of an elementary act of charge transfer reaction with regards to the energy spent on the transfer of charged reactants from water into the protein. The protein was treated as a structureless dielectric with a given optical and static dielectric constants surrounded by the aqueous phase. Reactions of different types (charge exchange between reactants, charge separation, neutralization, etc.) have been analyzed both under prevalence of purely electrostatic effects and under considerable nonelectrostatic contributions to the activation energies. It is shown that for all one-electron and most multi-electron reactions involving two reaction centres the energy spent for charged reactant transfer from water into protein is greater than the concomitant activation energy gain. The same effect takes place in a number of cases for multi-centre processes as well. To overcome the entropy hindrances, the reactants and catalysts must combine into multiparticle complexes, i.e. form microscopic regions of low dielectric constant. This results in increased effective activation energy as compared to reactions in water. It has been hypothesized that in order to make up for this loss the evolution has selected the proteins which are characterized by considerable intraglobular permanent electric fields. The presence in proteins of high concentrations of strongly polar peptide groups renders them advantageous in this respect over other polymers that are less polar.

Catalysis↗
PubMed1985

The negative cooperativity in cytochrome c oxidase redox reactions: the electrostatic effect.

The electrostatic interaction of two hemes is calculated on the basis of the data on the cytochrome c oxidase structure. The interaction energy corresponds to the positive shift by approximately greater than 100 mV of the redox potential of one of the hemes when the other is oxidized. This effect seems to be the most likely reason of the negative cooperativity in the redox behavior of cytochrome c oxidase.

Allosteric Regulation↗
PubMedDate not supplied

[Adsorption of dimethylsulfoxide on proteins].

A method for measuring the adsorption of dimethyl sulfoxide on native and denatured trypsin and albumin was developed. On native proteins, no positive adsorption was registered, and a slight negative adsorption within the limits of experimental error was observed. It was shown that the properties of denatured proteins depend on the mode and conditions of denaturation. On one of denatured trypsin specimens, positive adsorption of dimethyl sulfoxide was registered, on other specimens no adsorption was observed. The reason for this behavior lies in the hydrophobic nature of adsorption of dimethyl sulfoxide at the interface, while the surface of native protein globules and, probably, most denatured protein specimens is hydrophilic.

Adsorption↗