The method of tritium planigraphy, which provides comprehensive information on the accessible surface of macromolecules, allows an attempt at reconstructing the three-dimensional structure of a protein from the experimental data on residue accessibility for labeling. The semiempirical algorithm proposed for globular proteins involves (i) predicting theoretically the secondary structure elements (SSEs), (ii) experimentally determining the residue-accessibility profile by bombarding the whole protein with a beam of hot tritium atoms, (iii) generating the residue-accessibility profiles for isolated SSEs by computer simulation, (iv) locating the contacts between SSEs by collating the experimental and simulated accessibility profiles, and (v) assembling the SSEs into a compact model via these contact regions in accordance with certain rules. For sperm whale myoglobin, carp and pike parvalbumins, the lambda cro repressor, and hen egg lysozyme, this algorithm yields the most realistic models when SSEs are assembled sequentially from the amino to the carboxyl end of the protein chain.
Algorithms , Isotope Labeling/methods , Protein Structure, Tertiary , Tritium , Forecasting , Models, Molecular , Protein Structure, Secondary , Surface Properties
Classical model system: Poly-L-glutamic acid (Poly-Glu) was investigated in a disordered coil state (at pH-7.0) and in helix state (at pH 2.0) by Rayleigh scattering of Moessbauer radiation technique. Consider that the coil state of poly-Glu models unfolded (random coil) state and alpha-helix state models the fluctuating secondary structure (during consequent folding of protein) comparative analysis of dynamical properties of poly-Glu in different states with dynamical properties of different proteins in native state (alpha-helical myoglobin and HSA, partially beta-sheet lysozyme) and in intermediate (molten globule) state (alpha-lactalbumin) was performed. This comparison bring some surprising results: native alpha-helical proteins behave itself close to random coil, native partially beta-sheet protein behaves close to fluctuating secondary structure (alpha-helix) and the dynamic behaviour of molten globule state (partially beta-sheet alpha-lactalbumin) is not different from those behaviour of lysozyme and much more rigid than native alpha-helical proteins. As a result one cannot exclude the possibility that folding process and dynamical properties at different steps of the folding are very different for alpha-helical and beta-sheet proteins.
Globulins/chemistry , Polyglutamic Acid/chemistry , Protein Structure, Secondary , Spectroscopy, Mossbauer
The measurements of angle dependencies of total and elastic Rayleigh scattering of Mossbauer radiation intensities have been performed for human serum albumin (HSA) with hydration degrees h = 0.13 and h = 0.4. The extended model was developed for calculating the inelastic intensity of Rayleigh scattering. Original data for HSA and published data on met-Mb were fitted within the frame of this model. The best agreement with experiment was obtained when two types of intraglobular motions were taken into account: individual motions of small side-chain groups and cooperative (mechanical) motions of segments (most probable alpha-helices). Long-range correlated motions are essential at low hydration degree. The possibilities of application of the coherent version of RSMS technique are described.
Serum Albumin/chemistry , Humans , Metmyoglobin/chemistry , Models, Theoretical , Scattering, Radiation , Spectroscopy, Mossbauer
Murine tetrameric oxyhaemoglobin and insect monomeric erythrocruorin were studied. A doublet with the Lorentz form of lines (delta EQ = 2.22 mm s-1; delta alpha-Fe = 0.27 mm s-1; gamma 1/2 = 0.29 mm s-1) was observed in the oxyhaemoglobin spectrum at 4.2 K. In the 80-170 K interval the doublet components are distorted, but at T greater than 175 K, the lines again become symmetrical. In the 175-210 K region the value of gamma 1/2 is approximately 0.42 mm s-1. The profiles of the oxyhaemoglobin spectra are not dependent on the nature of the samples (blood; whole, or in aqueous or water-glycerol solutions at different pH values), or on the rate at which the latter are frozen. The oxyerythrocruorin spectra in aqueous solution and in a water-glycerol solution at 80 K and 170 K were doublets with Lorentz lines (the delta EQ values are equal to 2.20 mm s-1 and 2.12 mm s-1, respectively). It is concluded that the characteristics of the oxyhaemoglobin spectra reflect the specific electronic and structural properties of the oxy-complex in this protein. It was found that the oxyhaemoglobin spectra are very adequately described by two doublets with equal delta and gamma 1/2 values, but with different delta EQ values and relative intensities. A model is described in which these doublets correspond to two types of hydrogen bond associated with the distal histidine (E7), involving the terminal atom of molecular oxygen and the oxygen atom bound with the haem iron, respectively.
Insect Proteins , Oxyhemoglobins/chemistry , Animals , Insect Hormones/chemistry , Kinetics , Mathematics , Mice , Spectroscopy, Mossbauer/methods , Thermodynamics
Last experimental results of the study of protein dynamics by Mössbauer absorption spectroscopy and Rayleigh scattering of Mössbauer radiation are reviewed. Dynamical properties of proteins following from the theoretical treatment of these data are described.
Proteins , Humans , Light , Models, Theoretical , Protein Conformation , Scattering, Radiation , Spectrum Analysis
Hydration relationships of the elastic scattering fraction of Mössbauer radiation were studied for human serum albumin (HSA), pancreatic trypsin inhibitor and lysozyme within hydration degrees 0 less than or equal to h less than or equal to 0.75 g/g (at T = 295 degrees K) and temperatures 100K less than or equal to T less than or equal to 320 K (for HSA only at h = 0.03; 0.25; 0.41; 0.65). It is shown that the increase of both hydration degree above h greater than 0.1 and temperature above T greater than 200K leads to the appearance of intramolecular mobility in these proteins.
Proteins , Humans , Protein Conformation , Scattering, Radiation , Spectrum Analysis , Water
A review of the recent data on protein dynamics (mainly myoglobin) by X-ray technique, Mössbauer spectroscopy and Rayleigh scattering of Mössbauer radiation is given. The connection between dynamical and functional properties of biological systems are discussed.
Protein Conformation , Proteins/metabolism , Animals , Kinetics , Myoglobin/metabolism , Spectrum Analysis/methods , Temperature , X-Ray Diffraction/methods
Mobility of the Mössbauer label attached to the membrane proteins and the Mössbauer probe embedded into the lipid matrix of the bacterial chromatophores were studied. Positive correlation was established between the dynamic properties of hydrophobic compartments in the chromatophores and functional electron--transport activity at the level of quinone cofactors associated with the photosynthetic reaction centres.
Bacterial Chromatophores/ultrastructure , Intracellular Membranes/ultrastructure , Membrane Proteins/analysis , Rhodobacter sphaeroides/ultrastructure , Electron Transport , Spectrum Analysis/methods
The possible use of the method of 3H-label introduction by means of a 3H-atom beam for investigating the three-dimensional structure (surface) of proteins was studied. The intramolecular distribution of tritium in the N-terminal part of 3H-labelled sperm-whale myoglobin was studied. The results obtained are in good agreement with the X-ray analysis data on the polypeptide chain of the protein molecule. The possible use of this method for constructing precise three-dimensional models of polypeptides and proteins is discussed.
Myoglobin , Protein Conformation , Animals , Radioisotope Dilution Technique , Tritium , Whales , X-Ray Diffraction
Probabilities of the incorporation of tritium label into various amino acids with alanine as a standard have been determined by "bombing" solid targets with a 3H-atom beam (2 000 K). The results show that amino acids can be used with sufficient accuracy as models of amino acid residues in a polypeptide chain under 100%, accessibility. The resulting coefficients, if taken into consideration in the analysis of intramolecular distribution of tritium in short tryptic peptides of TMV protein, will yield an equiprobable distribution in the case when the label has been introduced into a peptide. The distribution is not uniform, however, if a labelled peptide has been isolated from an "irradiated" hydrolysate protein.
Amino Acids , Peptides , Protein Conformation , Proteins , Models, Biological , Molecular Weight , Peptide Fragments/analysis , Tobacco Mosaic Virus/analysis , Tritium , Trypsin , Viral Proteins
It has been shown that the mechanism of complex formation changes at variation of the detergent concentration. At low concentrations the complex is stabilized, mainly, by hydrophobic interactions. The boded molecules are immobilized inside the globules. At concentrations close to critical micellization concentration only some of the molecules take part in such strong binding, while the rest form weak bonds and may be removed by gel filtrations. Presumably in this case electrostatic interactions dominate in binding processes. At concentrations exceeding critical micellization concentration complex formation is controlled by electrostatic association of protein with aggregates of the detergent's molecules, stabilized, like micelles by hydrophobic forces. The complex is unstable.
Muramidase/metabolism , Sodium Dodecyl Sulfate , Kinetics , Protein Binding
A model was developed to describe conformational motion of macromolecules. A new formula was derived for the Debye-Waller factor in accordance with this model. The model proposes the existence of fluidlike motions of fragments within the volume of some A3. The temperature dependence of Rayliegh scattering of Mossbauer radiation was studied and calculated for chromatophores.
Molecular Conformation , Macromolecular Substances , Mathematics , Models, Biological , Spectrum Analysis/methods , Temperature
The temperature dependence of rayleigh scattering of Mössbauer radiation was studied for two samples of the chromatophores with different relative humidity (P/Ps = 0.35 and 0.94). New type of motion--the transitions between conformational substates was found above 200 degrees K. The model was developed to describe conformational motion in the membranes and membrane proteins. A new formula was derived for the Debye--Waller factor in accordance with this model. The model proposes the existence of fluid-like motions of protein fragments within the volume of some A3, restricted by the profile of the conformational potential. The correlation between decreasing of fR and increasing of electron transition rate was observed.
Bacterial Chromatophores , Rhodospirillum rubrum/ultrastructure , Bacterial Chromatophores/radiation effects , Chemical Phenomena , Chemistry , Cobalt Radioisotopes , Electron Transport , Mathematics , Models, Biological , Molecular Conformation , RNA, Transfer, Amino Acyl , Spectrometry, Gamma , Temperature
A new method of EPR-spectroscopy--the recombination of free radicals appearing as a result of indirect radiolysis of biological molecules after a low temperature irradiation--is applied to the study of molecular dynamics of phosphatidylcholine dimyristoyl in mass and in the structure of liposomes above and below the transition temperature. It was shown, that the mobility of lipid molecules in crystalline liposomes is higher than in the structure of liquid-crystalline liposomes. The addition of cholesterol in liposome membranes decreases the lateral molecular motion of lipids in crystalline and liquid-crystalline state, in the latter case the effect of cholesterol addition is more pronounced. The activation energy for the displacement of the fragments of lipid molecules and the lipid molecule as a whole was estimated, and it was shown, that lipid matrix possesses a high degree of heterogeneity.
Liposomes , Phosphatidylcholines , Dimyristoylphosphatidylcholine , Electron Spin Resonance Spectroscopy , Free Radicals , Freezing , Molecular Conformation , Spin Labels
