The role of local hydrogen bonds in formation of irregular regions of globular protein polypeptide chains]

An assumption is made on the substantial role of local hydrogen bonds in formation of irregular regions of globular protein polypeptide chains. The statistics of the amino acid composition of irregular regions is examined from this point of view. A statistical analysis of side group-backbone hydrogen bonds is carried out for three proteins: alpha-chy-motrypsin, lysozyme and myoglobin. It is shown that short side groups participate in formation of local hydrogen bonds more often than long ones. Conformations of amino acid residues in the first and the last positions are studied in beta-bends of 9 proteins. It is shown that over 70% of these residues are in conformations corresponding to the formation of local hydrogen bonds of three types: backbone-backbone, side groupbackbone, backbone-water molecule-backbone. Thus, the participation of the cooperative hydrogen-bonding network in stabilization of beta-bends is demonstrated.     

Proof of the formation of covalent bonds during staining of histologic preparations with procion dyes]

A method proving the formation of covalent bonds between a procion dye and histological slides is suggested based on the splitting of the covalent bound dye (I) into two parts, and on the synthesis of a new dye (II) on one of the parts of the dye (I) which was bound up with a covalent bond. If covalent bonds have not been formed while staining the slides a new dye (II) is not synthesized because two parts of the dye (I) have passed into solution.     

Determination of the time of polypeptide chain synthesis in animal experiments]

A method is worked out for the estimation of the time of polypeptide chain synthesis in animals in vivo. The time of the synthesis of "middle" polypeptide was found to be 1.45 min. in mouse liver cells under normal conditions, while in the presence of translation inhibitors, cycloheximide (20 mg/kg) and aurintricarboxilic acid (1 g/mg) the time of the synthesis increased in 2.7 and 2.5 times respectively. The time of polypeptide synthesis linearly increased with the increase of aurintricarboxilic acid dose.     

Double and bifurcated hydrogen bonds in alpha-helices of globular proteins]

The statistical analysis of hydrogen bonds distribution in space structures of globular proteins has been done. The parameters of H-bonds in the different secondary structures of globular proteins were collected. In alpha-helices besides the canonical 1-5 H-bonds (the mean length 3 A), 1-4 H-bonds were observed (the mean length 3.2 A). The histograms of length and angular distributions of the bonds are presented. It was found on the basis of quantum chemistry calculations that most H-bonds in alpha-helices are double or bifurcated.     

Formation of hydrogen bonds precedes the rate-limiting formation of persistent structure in the folding of ACBP

A burst phase in the early folding of the four-helix two-state folder protein acyl-coenzyme A binding protein (ACBP) has been detected using quenched-flow in combination with site-specific NMR-detected hydrogen exchange. Several of the burst phase structures coincide with a structure consisting of eight conserved hydrophobic residues at the interface between the two N and C-terminal helices. Previous mutation studies have shown that the formation of this structure is rate limiting for the final folding of ACBP. The burst phase structures observed in ACBP are different from the previously reported collapsed types of burst phase intermediates observed in the folding of other proteins.     

Effect of alcohols on polypeptide chain elongation and aminoacyl-tRNA formation.

The effect of alcohols (methanol, ethanol, and propanol) on polypeptide chain elongation was studied. In the E. coli and rat liver cell-free systems, the optimal concentration of Mg2+ decreased with increase of ethanol concentration, although the maximum polyphenylalanine synthesis decreased. Methanol had almost the same effect as ethanol. Propanol decreased the optimal magnesium concentration, but polyphenylalanine synthetic activity was markedly decreased. The shift of optimal Mg2+ concentration by ethanol was also observed in polylysine and polysome-dependent polypeptide syntheses. Even in the presence of spermidine, ethanol caused the shift of optimal Mg2+ concentration. Ribosome-bound Mg2+ was decreased by the addition of ethanol. A study of the effect of alcohols on aminoacyl-tRNA formation with ten amino acids in the absence of added Mg2+ showed that the formation of arginyl-, leucyl-, and valyl-tRNA was stimulated by the alcohols. Valyl-tRNA formation in the presence of alcohols was completely inhibited by EDTA, while that in the presence of Mg2+ was inhibited slightly by EDTA. No PP1-ATP exchange was observed when alcohol was used as the only stimulant of valyl-tRNA formation.     

Structure of collagen with a new net of hydrogen bonds]

A conformational variability of the collagen triple helix was studied with the methods of molecular mechanics. The Rich-Crick model with one hydrogen bond per tripeptide fragment or the model with two hydrogen bonds per tripeptide fragment were used for tripeptides forming the primary structure of the protein. Imino acid and amino acid residues were located in the second position of the tripeptide fragments in the first and second cases, respectively. Conformations on domain boundaries, which had alternating structures with one and two hydrogen bonds per tripeptide, were particularly studied. Essentially all types of collagen backbone composed of amino acid residues most frequently occurring in this protein were considered. A new model was suggested that combined elements of the Rich-Crick model and our new approach. This was shown to be stereochemically valid, energetically advantageous, and consistent with the experimental data. It was conclusively demonstrated that the primary structure of collagen determines its tertiary structure.     

Modelling hydrogen bonds in NN-dimethylformamide

N, N-dimethylformamide (DMF) is a ‘universal’ solvent with the simplest amide structure. DMF has different interactions with many polymers and biomolecules. It is therefore necessary to study systematically the interactions in DMF itself first. In this study, both FT-IR and two molecular theoretical methods (MP2 and DFT/B3LYP) were used to study various hydrogen bonding interactions in DMF molecules based on its weak H-bonding donors CH/CH₃ and strong H-bonding acceptor C = O. The possible H-bonding donors and acceptors in DMF molecules were first analysed followed by modelling the effect of different structural environments on vC = O bands in infrared spectra. Finally, H-bonding properties including distance, angles and the energy as well as the probability of H-bonding patterns were obtained. The results showed that there exist five possible different weak types of H-bonding dimers; among them, three dimers consist of a pair of weak H-bonds, whereas two other dimers have two pairs of H-bonds, leading to 14 (including eight different) H-bonds. Two types of dimers were dominant, whereas three others can be omitted.     

Sterospecific selection of nucleophilic compounds in the chymotrypsin-catalyzed formation of the peptide bonds]

The course of stereospecific selection of nucleophilic compounds was studied in the reaction of acyl-enzymes interaction with razemic substrate-like nucleophiles, e.g. amino acid esters, by measuring optical rotation or incorporation of labelled D-compounds. It was shown that the acyl-enzymes are not responsible for the stereospecific selection of substrate-like nucleophiles. Since stereospecific selection of nucleophiles occurs in some chymotrypsin-catalyzed reactions, such selection may be produced by chymotrypsin till the formation of an acyl-enzyme compound with the substrate at the enzyme-inhibitor stage (or the Michaelis complex) with nucleophilic compounds. Even under the optimal conditions no absolute stereospecific selection of nucleophiles occurred, as was observed in case of a substrate (a donor of the acyl amino acid residue), undergoing degradation. An essential role of a specific site of nucleophile binding in the reactions of chymotrypsin-catalyzed peptide bond formation, is emphasized.     

Interrelationship between glucoside-bound and free alcohols during ether oil formation in rose petals]

Some data on the dynamics of free and glucoside-bound monoterpenic and aromatic (beta-phenylethyl) ethers content and the changes in the beta-glucosidase activity in rose petals at different stages of the flower development and on the kinetics of enzymatic hydrolysis of these glucosides are presented. The phase specificity of beta-glucosidase coinciding with the maximal accumulation of glucoside-bound and free alcohols is revealed. The data obtained suggest that the formation of glucosides may precede the accumulation of corresponding free alcohols of terpenic and aromatic origin.     

Geometric criteria of hydrogen bonds in proteins and identification of "bifurcated" hydrogen bonds

Empirical criteria for identification of hydrogen bonds were analyzed to produce a set of geometrically consistent criteria. For a data set of 30 structures, application of a set of purely geometrical criteria, along with exclusion of abnormal backbone conformations, also excluded a common interaction of Ser/Thr side chains with Asp/Glu side chains ([ST]/[DE] pairs). These interactions were termed "bifurcated hydrogen bonds", which implies delocalization of a positively charged hydrogen of hydroxyl between the two acceptor atoms of the carboxylic group. These "bifurcated" interactions are among the most common packing patterns for [ST]/[DE] pairs of side chains. Therefore, the identification of hydrogen bonds cannot be based on geometrical criteria only and requires introduction of some physico-chemical criteria.     

Redistribution and loss of side chain entropy upon formation of a calmodulin-peptide complex

The response of the internal dynamics of calcium-saturated calmodulin to the formation of a complex with a peptide model of the calmodulin-binding domain of the smooth muscle myosin light chain kinase has been studied using NMR relaxation methods. The backbone of calmodulin is found to be unaffected by the binding of the domain, whereas the dynamics of side chains are significantly perturbed. The changes in dynamics are interpreted in terms of a heterogeneous partitioning between structure (enthalpy) and dynamics (entropy). These data provide a microscopic view of the residual entropy of a protein in two functional states and suggest extensive enthalpy/entropy exchange during the formation of a protein-protein interface.     

Modeling hydrogen bonds in three dimensions

The effective potential between two hydrogen bonded atoms is calculated on the basis of the Lippencott-Schroeder bent bond model, taken to be a typical model interaction. We differ from other calculations in that the minimum energy configuration for the proton is treated adiabatically, its position being recomputed at each value of the larger atoms separation. We find the typical hard core to have been a consequence of an artificial restriction of the proton to a fixed angle with the larger atom axis, basically a one-dimensional assumption. Free to move in three dimensions, the proton is squeezed off the axis as the separation narrows, and the hard core feature is gone. Depending on the degree of bond bending, the anharmonicity of the bond may be diminished, eliminated, or even reversed.     

Collapse of a polypeptide chain as a result of the intramolecular formation of antiparallel beta-sheets

The folded nature of an intramolecular antiparallel β-sheet suggests that the introduction of this structure into a statistical coil might be accompanied by a contraction of the chain. The magnitude of the contraction, and the conditions that produce the maximum contraction, have been assessed by the combination of generator matrices with an earlier formulation for the configuration partition function. The results show that the mean square dimensions do indeed pass through a minimum upon the transition from a statistical coil to an antiparallel β-sheet. The depth of the minimum is relatively insensitive to the values of δ and τ, provided the assignments are within the physically sensible range. (The borders of the ordered region are assumed to be of higher energy than the interior.) In contrast with the depth of the minimum, the β-sheet content that produces the minimum is quite sensitive to plausible variation in δ and τ. A much greater collapse of the chain can be produced by antiparallel β-sheet formation than that found in the course of the helix–coil transition. The collapse may cause the volume pervaded by the chain to come within an order of magnitude of the volume characteristic of the globular state.     

Rate of polymer formation and entropy production during competitive replication

Summary The rate of increase in the mean polymer formation rate constant during competitive replication byQ RNA variants (Kramer et al., 1974) has been shown to agree statistically with the variance in their formation rate constants. This result demonstrates that Fisher's fundamental theorem of natural selection (Fisher, 1930) can define time variations in the mean rate of synthesis for a heterogeneous population of replicating polymers. It was also revealed that RNA replication, far from equilibrium, accompanied a progressive decrease in the order of the entropy production derivative, with respect to time, that reached a maximum (with the next higher order being zero). Maximization of entropy at equilibrium, in compliance with the second law of thermodynamics, therefore appears as a natural extension of the earlier non-equilibrium pattern of entropy production within the system. The order of the zero-valued entropy production derivative was shown to be determined by the chemical affinity, and its rate of decrease was specified by the mean polymer formation rate constant.     

Formation and release of eukaryotic initiation factor 2 X GDP complex during eukaryotic ribosomal polypeptide chain initiation complex formation

The formation and release of an eukaryotic initiation factor (eIF)-2 X GDP binary complex during eIF-5-mediated assembly of an 80 S ribosomal polypeptide chain initiation complex have been studied by sucrose gradient centrifugation analysis. Isolated 40 S initiation complex reacts with eIF-5 and 60 S ribosomal subunits to form an 80 S ribosomal initiation complex with concomitant hydrolysis of an equimolar amount of bound GTP to GDP and Pi. Sucrose gradient analysis of reaction products revealed that GDP was released from ribosomes as an eIF-2 X GDP complex. Evidence is presented that eIF-5-mediated hydrolysis releases the GTP bound to the 40 S initiation complex as an intact eIF-2 X GDP complex rather than as free GDP and eIF-2 which subsequently recombine to form the binary complex. Furthermore, formation and release of eIF-2 X GDP from the ribosomal complex do not require concomitant formation of an 80 S initiation complex since both reactions occur efficiently when the 40 S initiation complex reacts with eIF-5 in the absence of 60 S ribosomal subunits. These results, along with the observation that the 40 S initiation complex formed with the nonhydrolyzable analogue of GTP, 5'-guanylylmethylene diphosphonate, can neither join a 60 S ribosomal subunit nor releases ribosome-bound eIF-2, suggest that following eIF-5-mediated hydrolysis of GTP bound to the 40 S initiation complex, both Pi and eIF-2 X GDP complex are released from ribosomes prior to the joining of 60 S ribosomal subunits to the 40 S initiation complex.