The structural principles of multidomain organization of the giant polypeptide chain of the muscle titin protein: SAXS/WAXS studies during the stretching of oriented titin fibres.

Elasticity of titin is a key parameter that determines the mechanical properties of muscle. These include reversibility, i.e., the muscle's capacity to change its length many-fold and return to its original state, and the transduction of passive tension generated by the stretched muscle. The morphology and elastic properties of oriented fibres of titin molecules were studied using SAXS and WAXS (small- and wide-angle X-ray scattering, respectively) and mechanical techniques. We succeeded in obtaining oriented filaments of purified titin suitable for diffraction measurements. Our X-ray data suggest a model of titin as a nanoscale, morphological, and aperiodical array of rigid Ig- and Fn3-type domains covalently connected by conformationally variable short loops. The line group symmetry of the model can be defined as SM with axial translation tau(infinity). Both tension transduction and high elasticity of titin can be explained in terms of crystalline polymer physics. Titin stretching experiments show that each individual titin macromolecule can adopt a novel two-phase state within the fibre. Conversion between high elasticity and strength can be explained as a phase transition under external tension. In the terms of the concept of orientational melting the origin of the functional heterogeneity along the titin strand becomes interpretable.

De novo design of fibrils made of short alpha-helical coiled coil peptides.

BACKGROUND: The alpha-helical coiled coil structures formed by 25-50 residues long peptides are recognized as one of Nature's favorite ways of creating an oligomerization motif. Known de novo designed and natural coiled coils use the lateral dimension for oligomerization but not the axial one. Previous attempts to design alpha-helical peptides with a potential for axial growth led to fibrous aggregates which have an unexpectedly big and irregular thickness. These facts encouraged us to design a coiled coil peptide which self-assembles into soluble oligomers with a fixed lateral dimension and whose alpha-helices associate in a staggered manner and trigger axial growth of the coiled coil. Designing the coiled coil with a large number of subunits, we also pursue the practical goal of obtaining a valuable scaffold for the construction of multivalent fusion proteins. RESULTS: The designed 34-residue peptide self-assembles into long fibrils at slightly acid pH and into spherical aggregates at neutral pH. The fibrillogenesis is completely reversible upon pH change. The fibrils were characterized using circular dichroism spectroscopy, sedimentation diffusion, electron microscopy, differential scanning calorimetry and X-ray fiber diffraction. The peptide was deliberately engineered to adopt the structure of a five-stranded coiled coil rope with adjacent alpha-helices, staggered along the fibril axis. As shown experimentally, the most likely structure matches the predicted five-stranded arrangement. CONCLUSIONS: The fact that the peptide assembles in an expected fibril arrangement demonstrates the credibility of our conception of design. The discovery of a short peptide with fibril-forming ability and stimulus-sensitive behavior opens new opportunities for a number of applications.

[The internal structure of soft and bony tissues studied by X-ray refraction introscopy]. / Issledovanie vnutrennei struktury miagkikh i kostnykh tkanei metodom rentgenovskoi refraktsionnoi introskopii.

Whether X-ray refractional introscopy can be used to examine biological objects was studied. The characteristic emission of an X-ray tube provided refractional radiograms of the rat heart and limb, which allow one to make out the details of the structure of soft tissues and bones invisible on absorption radiograms under the same conditions. High-contrast refractional images of calcified formations were seen in the real-time mode when they were registered with a two-coordinate TV detector. The potentialities of the new method and medical diagnostic means are discussed.

[Influence of low-intensity laser radiation on the formation of liquid crystalline structures in a solution of glycoproteins]. / Vliianie nizkointensivnogo lazernogo izlucheniia na formiravanie zhidkokristallicheskikh struktur v rastvore glikoproteinov.

Liquid-crystalline structure formation in glycoprotein solutions irradiated by helium-neon laser in the presence of hydrogen peroxide was observed by both polarizing microscopy and spectrophotometry. High molecular weight (2.10(6) Da) and heavily glycosylated (about 80%) glycoprotein was isolated from the mucus layer of pig small intestine. Remarkable changes of both optic parameters of the solutions and the morphology of liquid-crystalline structures were detected in irradiated samples compared to the non-irradiated ones.

[Study of the structure of metal-binding centers of calmodulin by EXAFS-spectroscopy]. / Issledovanie struktury metallsviazyvaiushchikh tsentrov kal'modulina metodom EXAFS-spektroskopii.

An investigation of Ca2+-binding centers of calmodulin was carried out by EXAFS-spectroscopy. The experimental results for protein preparations of calmodulin in which Ca2+ was isomorphically replaced by Tb3+ were obtained by a spectrometer working at the Institute of Nuclear Physics. For spectra analyses a standard method of Fourier transformation was used. Coincidence main maxima on phi (r) curves and identity of Fourier transformation for calmodulin and parvalbumin in the 2-6 A interval allow to infer the identity of Ca2+-binding centers of calmodulin and parvalbumin.

[Interpretation of the diffraction picture of a skeletal muscle during the active phase of contraction]. / K interpretatsii difraktsionnoi kartiny skeletnoi myshtsy v aktivnoi faze sokrashcheniia.

New data concerning changes in the diffraction patterns at muscle activation, obtained by X-ray studies with a high resolution rate helped to interpret the diffraction patterns of the skeletal muscle in the active phase of contraction. Changes in the intensity of the meridianal reflex 143 A at the time of isomeric contractions and during rapid mechanical muscle stimulus are discussed. Experimental data analysis and calculations of the diffraction pattern, corresponding to the states of rest and contraction, showed, that the observed changes can be explained by the model of contactless interaction of myosin bridges with fine fibers. The diffraction pattern at the active phase of contraction showed that the bridges are near the fine fiber, but are not attached to the specific centers of binding on the actin globules.

[Study of Ca2+-binding centers of parvalbumin by the distant fine structure of x-ray absorption spectra]. / Issledovanie Ca2+-sviazyvaiushchikh tsentrov parval'bumina po dal'nei tonkoi strukture rentgenovskikh spektrov pogloshcheniia.

An investigation of Ca2+-binding centers of parvalbumin II and III by analysing distant fine structure of X-ray absorption spectra of metal was performed. Protein preparations of parvalbumin II and III in which Ca2+ was isomorphically replaced by Tb3+ were studied. For spectra analyses a standard method of Fourier transformation was used. The middle of the first absorption maximum was taken as origin for energy calculations. Comparison of spectra and modules of Fourier transformations for normalized oscillations of the X-ray spectra of absorption of the II and III components, revealed that the spectra and Fourier-transformants coincide in the 2--6 A interval. This allows to infer the coincidence of the coordinate numbers, average interatomic distances and their dispersions in Ca2+-binding centers of the two protein components.

[High time resolution x-ray study of the dynamics of a single muscle contraction]. / Rentgenograficheskoe issledovanie dinamiki odinochnogo myshechnogo sokrashcheniia s vysokim vremennym razresheniem.

A method of the diffraction cinema which enables to study the time-course of structural changes during twitch contraction is described. The method is based on using synchrotron radiation, position-sensitive counter and small-angle focusing X-ray camera. Only 0.1 s is required to record a good muscle X-ray diagram: meridional diagram contains all layer-lines beginning with the 429 A; the equatorial diagram contains 5 reflections including very weak alpha-reflection. The method allows to record 64 sequent diffraction patterns with different duration (1--2000 ms). The experiment is handled by a computer. Some tens of the films of isometric twitch contraction with time resolution of 3--20 ms have been obtained. During isometric contraction considerable changes in the intensity of both meridional and equatorial reflections were found. The changes were interpreted as indicating movement of cross-bridges toward the thin filaments. During the latent phase there are no visible changes in the intensity of the reflections; the result indicates that during this phase there are no structural changes in position and configuration of cross-bridges.

[Small-angle diffuse x-ray scattering on fine muscle filaments]. / Malouglovoe diffuznoe rasseianie rentgenovykh luchei na tonkikh nitiakh myshtsy.

Theoretical small-angle diffuse scattering curves from muscle thin filament models have been calculated. The curves reveal a maximum at 115' scattering angle. It has been shown that the maximum is due to the pitch of F-actin helix. Theoretical curves are in good agreement with the earlier obtained curves of small-angle diffuse scattering from F-actin dilute solutions.

[Use of linear proportional chambers for small-angle synchrotron radiation scattering in solutions of biopolymers]. / Ispol'zovanie odnokoordinatnykh proportsional'nykh kamer v metode malouglovogo rasseianiia sinkhrotropnogo izlucheniia na rastvorakh biopolimerov

The results of applying linear position sensitive proportional counters to diffuse small-angle synchrotron radiation scattering to biopolymer solutions are presented. Synchrotron radiation of the VEPP-3 storage ring (Institute of Nuclear Physics, Novosibirsk) was used. On the basis of small-angle scattering curve obtained from protein pepsinogen it has been shown that the exposure is about 100 times less than with conventional X-ray technique.

[Experience in the use of synchrotron radiation for the x-ray study of biopolymers]. / Opyt ispol'zovaniia sinkhrotronnogo izlucheniia dlia rentgenograficheskogo issledovaniia biopolimerov

The results of methodical work, carried out on the sources of synchrotron radiation (SR) with the aim of using SR as a powerful source of X-rays for studying biopolymer structure, are presented. The questions of monochromatization are considered. The technique designed for photoregistration of diffraction patterns within the wide range of scattering angles is described. X-ray diffraction patterns of feather ceratin, collagen and striated muscle are obtained with exposure periods ten times less than those in the case of X-ray tubes. The high resolution of diffraction lines, the absence of parasitic phone and the presence of reflections within the wide range of scattering angles are characteristic of these patterns.