Biomedical subjects

I Roterman

Publications and source records attributed to I Roterman.

At least 19 recordsLinked to original sources

PubMed2004

Limited conformational space for early-stage protein folding simulation.

MOTIVATION: The problem of early-stage protein folding is critical for protein structure prediction. The model presented introduces a common definition of protein structures which may be treated as the possible in silico early-stage form of the polypeptide chain. Limitation of the conformational space to the ellipse path on the Ramachandran map was tested as a possible sub-space to represent the early-stage structure for simulation of protein folding. The proposed conformational sub-space was developed on the basis of the backbone conformation, with side-chain interactions excluded. RESULTS: The ellipse-path-limited conformation of BPTI was created using the criterion of shortest distance between Phi, Psi angles in native form of protein and the Phi, Psi angles belonging to the ellipse. No knots were observed in the structure created according to ellipse-path conformational sub-space. The energy minimization procedure applied to ellipse-path derived conformation directed structural changes toward the native form of the protein with SS-bonds system introduced to the procedure. AVAILABILITY: Program 'Ellipse' to create the ellipse-path derived structure available on request: myroterm@cyf-kr.edu.pl

Algorithms↗

PubMed2004

Lysozyme folded in silico according to the limited conformational sub-space.

The conformational sub-space oriented on early-stage protein folding is applied to lysozyme folding. The part of the Ramachandran map distinguished on the basis of a geometrical model of the polypeptide chain limited to the mutual orientation of the peptide bond planes is shown to deliver the initial structure of the polypeptide for the energy minimization procedure in the ab initio model of protein folding prediction. Two forms of energy minimization and molecular dynamics simulation procedures were applied to the assumed early-stage protein folding of lysozyme. One of them included the disulphide bond system and the other excluded it. The post-energy-minimization and post-dynamics structures were compared using RMS-D and non-bonding contact maps to estimate the degree of approach to the native, target structure of the protein molecule obtained using the limited conformational sub-space for the early stage of folding.

Computer Simulation↗

PubMed2004

Intramolecular signaling in immunoglobulins -- new evidence emerging from the use of supramolecular protein ligands.

The postulated intramolecular signaling in immunoglobulins generated by antigen binding has been controversial for years. The high heterogeneity of immune complexes as signaling systems and the requirement of the immobilized antigen form for efficient triggering of effector activity is likely the reason for the lack of clarity. Here we present new evidence supporting the notion of intramolecular signaling, based on the use of supramolecular dyes that bind to signal-derived specific sites in immunoglobulins.

Antigen-Antibody Complex↗

PubMed2002

Egg yolk platelet proteins from Xenopus laevis are amyloidogenic.

The association of amphibian (Xenopus laevis) egg yolk platelet proteins, represented predominantly by lipovitellin, was studied as a model of the formation of amyloid deposits. Two kinds of molecular organization formed by this protein material - native and heat-denatured - were found to exhibit amyloid properties although they differ significantly in structural organization. The first consisted in protein molecules arranged in the natural, physiological, net-like platelet organization, with a tendency to orient uni-directionally. The second was obtained by the gradual removal of Congo red from lipovitellin denatured by heating in an excess of dye. This procedure produced the twisted fibrillar organization of molecules typical for amyloids, represented predominantly by end-to-end associated major polypeptide chains of lipovitellin. Both native and denatured structural forms bind Congo red and produce a green birefringence effect, confirming the near parallel alignment of the complexed Congo red molecules. However, a dye(1,4-bis(1-amino-4-sulfonaphtyl-2-azo)phenylene) closely related to Congo red but with a very weak self-assembling tendency appeared inactive when the spectral shift was studied in a cross-polarization system, indicating in this way that dye supramolecularity is an extra factor which may determine binding to amyloid proteins and specific spectral effects.

Amyloid↗

PubMed2001

Heat-induced formation of a specific binding site for self-assembled Congo Red in the V domain of immunoglobulin L chain lambda.

Moderate heating (40-50 degrees C) of immunoglobulins makes them accessible for binding with Congo Red and some related highly associated dyes. The binding is specific and involves supramolecular dye ligands presenting ribbon-like micellar bodies. The L chain lambda dimer, which upon heating disclosed the same binding requirement with respect to supramolecular dye ligands, was used in this work to identify the site of their attachment. Two clearly defined dye-protein (L lambda chain) complexes arise upon heating, here called complex I and complex II. The first is formed at low temperatures (up to 40-45 degrees C) and hence by a still native protein, while the formation of the second one is associated with domain melting above 55 degrees C. They contain 4 and 8 dye molecules bound per L chain monomer, respectively. Complex I also forms efficiently at high dye concentration even at ambient temperature. Complex I and its formation was the object of the present studies. Three structural events that could make the protein accessible to penetration by the large dye ligand were considered to occur in L chains upon heating: local polypeptide chain destabilization, VL-VL domain incoherence, and protein melting. Of these three possibilities, local low-energy structural alteration was found to correlate best with the formation of complex I. It was identified as decreased packing stability of the N-terminal polypeptide chain fragment, which as a result made the V domain accessible for dye penetration. The 19-amino acid N-terminal fragment becomes susceptible to proteolytic cleavage after being replaced by the dye at its packing locus. Its splitting from the dye-protein complex was proved by amino acid sequence analysis. The emptied packing locus, which becomes the site that holds the dye, is bordered by strands of amino acids numbered 74-80 and 105-110, as shown by model analysis. The character of the temperature-induced local polypeptide chain destabilization and its possible role in intramolecular antibody signaling is discussed.

Amino Acid Sequence↗

PubMed2001

Evidence that supramolecular Congo red is the sole ligation form of this dye for L chain lambda derived amyloid proteins.

The mechanism of Congo red binding to amyloid protein was studied in order to establish which of two structural dye versions present in water solutions--unimolecular and supramolecular--represent its actual ligation form. Immunoglobulin L chain lambda of amyloidogenic nature, expressed by Congo red binding and easy gel formation, was used as the model amyloid protein. Congo red was coassembled with rhodamine B, designed to be a marker of the Congo red micellar organisation in complexation with protein. The particular suitability of rhodamine B for this role results from significant difference in its binding affinity to Congo red and to protein. It associates readily with Congo red, becoming incorporated into its micellar organisation, but as homogenous dye it shows an almost complete inability to bind to protein. In view of these properties, Congo red was used as a vehicle to draw rhodamine B into complexation with protein, at the same time supplying evidence of its supramolecular ligation form. The results show that both soluble amyloid precursor L chain and the derived gel material attach rhodamine B coassembled with Congo red but not the homogenous rhodamine B. Despite its dynamic, supramolecular character, Congo red participates in complexation with amyloid proteins as an integral ligand unit.

Amyloid beta-Protein Precursor↗

PubMed1999

Supramolecularity creates nonstandard protein ligands.

Congo red and a group of structurally related dyes long used to stain amyloid proteins are known to associate in water solutions. The self-association of some dyes belonging to this group appears particularly strong. In water solutions their molecules are arranged in ribbon-like micellar forms with liquid crystalline properties. These compounds have recently been found to form complexes with some native proteins in a non-standard way. Gaps formed by the local distribution of beta-sheets in proteins probably represent the receptor sites for these dye ligands. They may result from higher structural instability in unfolding conditions, but also may appear as long range cooperative fluctuations generated by ligand binding. Immunoglobulins G were chosen as model binding proteins to check the mechanism of binding of these dyes. The sites of structural changes generated by antigen binding in antibodies, believed to act as a signal propagated to distant parts of the molecule, were assumed to be suitable sites for the complexation of liquid-crystalline dyes. This assumption was confirmed by proving that antibodies engaged in immune complexation really do bind these dyes; as expected, this binding affects their function by significantly enhancing antigen binding and simultaneously inhibiting C1q attachment. Binding of these supramolecular dyes by some other native proteins including serpins and their natural complexes was also shown. The strict dependence of the ligation properties on strong self-assembling and the particular arrangement of dye molecules indicate that supramolecularity is the feature that creates non-standard protein ligands, with potential uses in medicine and experimental science.

Animals↗

PubMed1998

Supramolecular ligands: monomer structure and protein ligation capability.

The aim of this work was to define the chemical structure of compounds self-assembling in water solutions, which appear to interact with proteins as single ligands with their supramolecular nature preserved. For this purpose the ligation to proteins of bis azo dyes, represented by Congo red and its derivatives with designed structural alterations, were tested. The three parameters which characterize the reactivity of supramolecular material were determined in the same conditions for all studied dyes. These were: A) stability of the assembly products; B) binding to heat-denatured protein (human IgG); and C) binding to native protein (rabbit antibodies in the immune complex) measured by the enhancement of hemagglutination. The structural differences between the Congo red derivatives concerned the symmetry of the molecule and the structure of its non-polar component, which occupies the central part of the dye molecule and is thought to be crucial for self-assembly. Other dyes were also studied for the same purpose: Evans blue and Trypan blue, bis-ANS and ANS, as well as a group of compounds with a structural design unlike that of bis azo dyes. Compounds with rigid elongated symmetric molecules with a large non-polar middle fragment are expected to form a ribbon-like supramolecular organization in assembling. They appeared to have ligation properties related to their self-assembling tendency. The compounds with different structures, not corresponding to bis azo dyes, did not reveal ligation capability, at least in respect to native protein. The conditions of binding to denatured proteins seem less restrictive than the conditions of binding to native molecules. The molten hydrophobic protein interior becomes a new binding area allowing for complexation of even non-assembled molecules.

Congo Red↗

PubMed1997

Effect of self association of bis-ANS and bis-azo dyes on protein binding.

A correlation was found between the ability of dyes (ANS, bis-ANS, Congo red, Evans blue) to form self-associated supramolecular structures in water and their tendency to form complexes with proteins. The self-association ability of dyes was measured as the resistance of a molecular sieve to their penetration. Quantitative evaluation of dye-protein interaction involved measuring the effect of dye on antibodies that agglutinate sheep red blood cells. Enhancement of agglutination by dye was assumed to represent its protein complexation ability. The results confirm that, relative to monomers, self-associated ligands also have altered protein binding properties.

Anilino Naphthalenesulfonates↗

PubMed1997

The use of congo red as a lyotropic liquid crystal to carry stains in a model immunotargeting system--microscopic studies.

The lyotropic liquid crystal dye-Congo Red was used as a carrier in a model immunotargeting system constructed from sheep red blood cells (SRBC) representing the antigen target and rabbit IgG anti-SRBC as the specific driving immunoglobulin. Rhodamine B and Hemin stains were chosen as example chemicals carried to the target. The carried stains were introduced to the micellar organization of Congo Red by intercalation. Preserving its supramolecular organization, Congo Red binds spontaneously and selectively to antibodies that have altered structure extorted by interaction with the antigen in the immune complex. The functionality of the studied immunotargeting model was verified by fluorescence and electron microscopy. The results indicate that the supramolecular nature of protein ligands offers new ligation capabilities possibly useful for carrying stains or drugs in immune-oriented systems.

Animals↗

PubMed1996

Congo red-stabilized intermediates in the lambda light chain transition from native to molten state.

Disruption of tertiary interaction makes a protein accessible to penetration by different small molecular compounds. Their interaction may stabilize the altered protein conformation. Congo red is proposed here as a stabilizer of the molten globule state and also of highly reversible intermediates in the transition from native to molten state. Human immunoglobulin lambda light chain (dimer) was used. Two protein-Congo red complexes were found after heating lambda chain in the presence of Congo red. They differed in the amount of attached dye molecules. The binding of dye was interpreted as a two-step dye penetration process involving the peripheral parts of the protein in the first step (at lower temperatures). It was concluded that the liquid crystal properties of Congo red enable it to form specific complexes with proteins, which have become accessible to penetration by ligands after global or local disruption of tertiary interaction. This dye may thus be used as a stabilizer of unfolding intermediates in the step preceding the molten globule state.

Anilino Naphthalenesulfonates↗

PubMed1995

Modelling the optimal simulation path in the peptide chain folding--studies based on geometry of alanine heptapeptide.

The theoretical background of a simple model of polypeptide chain structure using two parameters: R (A)--the radius of curvature for each pentapeptide chain fragment in the protein, and V (deg)--the dihedral angle between two consecutive peptide bond planes, is presented. The mathematical relationship between these two geometrical parameters leads to the optimal searching path for low-energy peptide conformations. This R versus V relation, corresponding to low-energy structures in Ramachandran plot, appeared to fit the square function well. Here, the minimum of this function is taken as the optimal starting point for the minimization of all second-order conformations in the peptide chain. The extension, including all structures that satisfy the square function between V and R, showed the Phi, Psi angles that are optimal in searching for the path to low-energy structures. The path is an ellipse connecting the alpha R-, beta- and alpha L-structures, indicating the possible transitions from one to the next.

Alanine↗

PubMed1995

Geometrical analysis of structural changes in immunoglobulin domains' transition from native to molten state.

Molecular dynamics simulation (300, 320, 340 K) performed on the Fab (Kol) fragment of immunoglobulin G revealed that the structural changes associated with relaxation of peptides after their release from the stabilized by the tertiary interaction native conformation may be considered characteristic of the transition from native to molten state. The configuration of peptide chains at temperatures close to melting, liberated from the constraints associated with tertiary packing, was found to deviate toward helical rather than extended forms. The direction of the shift is diagonal on the phi-psi map. The torsional angles tend to concentrate in the Cea7 region, and some leak to the alphaR area. The geometrical parameters designed to describe the configuration of the peptide chain in Fab fragment also confirmed that during melting the peptides generally moved toward helical form.

Crystallography, X-Ray↗

PubMed1995

The geometrical analysis of peptide backbone structure and its local deformations.

A proposal to use a unified description of protein structures is presented, assuming that all peptide chain conformations (including beta-structure) may be considered as helix-derived. In this understanding the beta-conformation of the peptide chain may be obtained by extending or unwinding the helical form. A unified description could be based on a commonly defined internal axis (Z-axis) determined by the averaged direction of the CO bonds in a pentapeptide analytical unit. Five parameters were proposed to describe the peptide conformation: i) radius of curvature (R), calculated from the position of five consecutive peptide bond planes; ii) horizontal angular displacement (H) of the neighboring (i+1-th) versus the central amino acid residue (i-th) in the pentapeptide fragment; iii) vertical angular displacement (Vi) of the peptide bond plane of the i-th residue versus the Z-axis; iv) vertical angular displacement (Vi+1) of the i+1-th peptide plane measured versus the Z-axis; and v) vertical rise (VR) defined for the i+2-th residue versus the i-th residue. This geometrical analysis was used to study the mechanism of a possible transition from alpha R to beta and to alpha L transition, based on data taken from the known Phi, Psi distribution in proteins. This analysis was also designed and preliminarily used to reveal local peptide backbone distortions and their distributions in proteins for structure-function relation studies.

Animals↗

PubMed1995

The detection of specific acute phase serum protein complexes and immune complexes by congo red binding.

The study concerns the problem of selective binding of Congo Red dye to serum proteins. The dye-binding structural motifs were assumed to appear in proteins as the result of local disruptions of tertiary interaction associated with biological activities. Functional structures of serum proteinase inhibitors and haptoglobins induced by interaction of these proteins with their natural substrates were found to bind Congo Red. Soluble immune complexes formed by bivalent hapten-bridged anti-TNP rabbit antibodies also appeared able to interact with the dye. The possible use of these effects for diagnostic analysis was suggested.

Acute-Phase Proteins↗

PubMed1994

Heat-induced structural changes in the Fab fragment of IgG recognized by molecular dynamics stimulation--implications for signal transduction in antibodies.

Molecular dynamics simulation was used to identify the conformational alterations in the Fab fragment (Kol), driven by heating at 300, 320, and 340 K. Comparison of heat-modified VH,CH1 and VL,CL domain structures with the corresponding crystal conformations revealed specific differences, most definitely expressed in the CH1 domain. These are dislocations of predominantly peripheral peptide loops exposed to the V-C interdomain interface, comprising in particular the 175-185 amino acids of the heavy chain, as well as the 112-123 amino acids of the interdomain hinge fragment. The deviations, limited to peripheral domain regions at 300 and 320 K, spread at 340 K. The resulting relaxation of the tertiary packing in the protein (including the hydrophobic core) initiates global melting of the domain. These theoretical results were supported by experimental findings concerning penetration and binding of dyes (Congo Red, Trypan Blue, ANS) to the protein. Packing relaxation of the CH1 domain is turned on after disruption of the specific peptide arrangement formed at the V-C interdomain interface basically at the hinge portion (117-122) and at fragments of adjacent peptide loops (149-154, 171-179) of CH1 origin, most probably playing the role of a switching mechanism. The dislocations also comprise the 131-141 amino acids of the loops accommodated at the CH1-CH2 interface. However, the lack of crystallographic data concerning the Fab-Fc interface limits discussion of this effect to speculation. It was concluded that the unconcerted movements of the V and C parts of the Fab fragment are an intrinsic driving mechanism, introducing structural alterations into the C domains. It is suggested that the domain relaxation, induced by heating or mechanical constraints, allows for intermolecular interactions, affecting in this way the stability of the immune complex.

Computer Simulation↗

PubMed1994

Bis azo dye liquid crystalline micelles as possible drug carriers in immunotargeting technique.

Lyotropic liquid crystal mesophases, represented in this work by Congo Red bis azo dye solution, were proposed as systems to carry drugs to immuno-selected targets. The possible use of Congo Red for this aim arises from its liquid crystalline features, enabling it to attach to immune complexes as polymolecular, ordered conglomerates and simultaneously to incorporate many organic compounds into its mesophase, largely independent of the water with a specific but adaptive molecular organization. Molecules with planar rigid structure, and/or large hydrophobic fragments, especially those with a positively charged group in the molecule, are found to be incorporated best. Rhodamine B, Rhodamine 6G and adriamycine, which have the assumed binding features, were tested as model compounds and were found to be readily engaged into Congo Red mesophase. The effect of hydrophobicity on ligand binding was evaluated following the incorporation of homologic 10, 12, 14, 16 carbon chain organic acids and the effect of charge using small mobile tandem molecules of pI differing by a few pH units (lysine-norleucine, tyrosine-tyramine). Positive charge seems to affect binding especially by influencing the organization of molecules and the shape of the micelle simultaneously. Congo Red immobilized to heat-aggregated immunoglobulins and antibodies in the immune complex was found to retain its binding ability, confirming its possible usefulness for drug transport.

Antibodies↗

PubMed1994

The melting of native domain structure in effector activation of IgG studied by using congo red as a specific probe.

The nature of structural changes in IgG molecules associated with the binding to antigen and/or heat aggregation was studied using bis azo dye (Congo Red) as the specific probe. It was found, that protein conformation responsible for binding the dye represents an unfolding intermediate with properties corresponding to a molten globule state. The properties of the dye-protein complex reveal the signs of an unfolding of the peptide chain with simultaneously preserved relatively compact packing. Immunoglobulins which were induced by heating, or binding to antigen in order to form the complex with dye ligands, become more susceptible for digestion. The main peptide of molecular weight 30,000 D which appears in products was suggested to originate from a heavy chain after its splitting in the region of CH1 domain. The energetic evaluation of stability of IgG domains also indicates that CH1 is the least stable fragment of the heavy chain and its conformation may be destabilized first. It was concluded that destabilized tertiary packing of antibodies bound to antigen may favour the association of closely situated immunoglobulin molecules increasing the stability of the immune complex and influencing in the result its effector activity.

Antigen-Antibody Complex↗