Discovery Studio 2.0的模块扩展及应用:残基水平非键相互作用能量的自动批量计算

非键相互作用对于生物体系中的分子识别和结合过程起着关键作用。然而,传统的方法并不能在残基水平自动批量计算非键相互作用。近年来,已经发展了一些方法和工具进行非键相互作用的计算分析。该文研究发展了一种可以自动计算残基间非键相互作用的方法,即用Perl脚本调用Discovery Studio 2.0(DS 2.0,Accelrys Inc.)底层模块中的非键相互作用协议,实现了直接利用命令行批量计算非键相互作用能量,而无需通过DS2.0的图形界面。该方法扩展了DS2.0的计算模块,并于近期运用到了复合结构的研究分析中。     

Total free energy analysis of fully hydrated proteins

The total free energy of a hydrated biomolecule and its corresponding decomposition of energy and entropy provides detailed information about regions of thermodynamic stability or instability. The free energies of four hydrated globular proteins with different net charges are calculated from a molecular dynamics simulation, with the energy coming from the system Hamiltonian and entropy using multiscale cell correlation. Water is found to be most stable around anionic residues, intermediate around cationic and polar residues, and least stable near hydrophobic residues, especially when more buried, with stability displaying moderate entropy-enthalpy compensation. Conversely, anionic residues in the proteins are energetically destabilized relative to singly solvated amino acids, while trends for other residues are less clear-cut. Almost all residues lose intraresidue entropy when in the protein, enthalpy changes are negative on average but may be positive or negative, and the resulting overall stability is moderate for some proteins and negligible for others. The free energy of water around single amino acids is found to closely match existing hydrophobicity scales. Regarding the effect of secondary structure, water is slightly more stable around loops, of intermediate stability around $\beta$ strands and turns, and least stable around helices. An interesting asymmetry observed is that cationic residues stabilize a residue when bonded to its N-terminal side but destabilize it when on the C-terminal side, with a weaker reversed trend for anionic residues.     

An analysis of side-chain conformation in proteins.

The crystal structures of a number of globular proteins are currently available. An analysis of the distribution of side-chains among different allowed conformations in these proteins has been carried out. The observed conformations of individual residues are discussed on the basis of well-known stereochemical criteria. The population distribution of side-chains in different allowed regions in conformational space can be explained largely on the basis of simple steric considerations. In addition to examining the conformational behaviour of individual residues, some population distributions of conformational angles of general interest involving groups of residues have also been analyzed.     

Some chemical and structural consequences of non-bonded interactions

Unusual geometries at first row atoms, and maximum coordination numbers of first and second row atoms are discussed in terms of non-bonded interactions. Structures of closed shell molecules are divided into three categories in which respectively inter-electron repulsions dominate, electronic and nuclear terms are in balance, and inter-nuclear repulsions dominate.     

Structure-activity relationships in engineered proteins: analysis of use of binding energy by linear free energy relationships

The activity of mutant enzymes can be analyzed quantitatively by structure-activity relationships in a manner analogous to Br?nsted or Hammett plots for simple organic reactions. The slopes of such plots, the beta values, indicate for the enzymatic reactions the fraction of the overall binding energy used in stabilizing particular complexes. In particular, information can be derived about the interactions between the enzyme and the transition state. The activities of many mutant tyrosyl-tRNA synthetases fit well simple linear free energy relationships. The formation of enzyme-bound tyrosyl adenylate (E.Tyr-AMP) from enzyme-bound tyrosine and ATP (E.Tyr-ATP) results in an increase in binding energy between the enzyme and the side chain of tyrosine and the ribose ring of ATP. Linear free energy plots of enzymes mutated in these positions give the fraction of the binding energy change that occurs on formation of the transition state for the chemical reaction and the various complexes. It is shown that groups that specifically stabilize the transition state of the reaction are characterized by beta values much greater than 1. This is found for residues that bind the gamma-phosphate of ATP (Thr-40 and His-45) and have previously been postulated to be involved in transition-state stabilization.(ABSTRACT TRUNCATED AT 250 WORDS)     

Modulation of diorganoyl dichalcogenides reactivity by non-bonded nitrogen interactions

The study was designed to explore the biochemical influence of non bonding nitrogen interactions (N?Se/S) on organochalcogens potency. Approximately five and six times higher thiol peroxidase (TPx) like activity was observed for compound (C)-2 than C-1 and C-3, respectively. C-2 also displayed significantly (p < 0.05) higher activity in 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging and deoxyribose degradation assays. All compounds, except C-4 and C-6 significantly inhibited Fe (II) and sodium nitroprusside (SNP) induced thiobarbituric acid reactive species (TBARS) production in rat’s brain, liver and kidney preparations with highest activity observed for C-2. The highest C-2 activity was attributed to the presence of non-bonded nitrogen interactions which were absent in C-1 and blocked with butoxycarbonyl (BOC group) in C-3. The same structural activity analogy was extended to organosulfur compounds and it was observed that compound with non-bonding nitrogen interactions, i.e. C-5 has significantly (p < 0.05) higher TPx like activity than C-6 and C-4. C-5 at the highest tested concentration significantly (p < 0.05) protected against Fe (II) and SNP induced TBARS formation in rat’s brain, kidney and liver preparations but did not display activity in DPPH and deoxyribose degradation assays. This study confirms the influence of not only N?Se interaction but also for the first time the effect of non bonded N?S interactions on organochalcogens potency. C-2 (with the highest activity) was also tested in vivo and was administered at three different doses, i.e. 15, 30 and 50 mg/kg to get an exact idea about its interaction with thiol containing molecules (NPSH) and enzyme α-ALA-D (sulfhydryl containing enzyme). Oxidative stress parameters, i.e. free radical concentration by dichlorofluoreseein (DCF) assay, TBARS, ascorbic acid level, hepatic (ALT and AST) and renal (urea and creatinine) toxicity markers were also estimated to get an insight about its possible toxicological profile. Our data indicates that C-2 has higher TPx and Antioxidant activity and importantly, C2 did not induce toxicity even when tested at relatively high doses, indicating that its pharmacological properties should be further explored in models of diseases associated with oxidative stress.     

Navigation and analysis of the energy landscape of small proteins using the activation-relaxation technique

The resolution of the protein folding problem has been tied to the development of a detailed understanding of the configurational energy or of the free energy landscape associated with these molecules. Using the activation-relaxation technique and a simplified energy model, we present here a detailed analysis of the energy landscape of 16-residue peptide that folds into a beta-hairpin. Our results support the concept of an energy landscape with an effective topology consistent with a scale-free network.     

An electrophoretic analysis of the basic nuclear proteins of ram spermatids

By a two-step separation procedure, round and elongated spermatid populations were obtained with purities of 90–97% and an intermediate spermatid population with a purity of 85%. Cells of ram testes were first separated at unit gravity and 5 spermatid fractions were collected. They were centrifuged for 23 min at 4 000 g in linear and non-linear isotonic polymer-free colloidal silica gradients. Improvements were made in centrifuge tubes and in gradient collection for optimum results. Fractions were analysed using a phase contrast microscope for cell counting and determination of cell viability. True cell density was measured and found to increase during late spermiogenesis from 1.045 g/cm³ for round spermatids to 1.152 g/cm³ for testicular spermatozoa, thus reflecting an increase in the nuclear density.     

An X-ray diffraction analysis of oriented lipid multilayers containing basic proteins

X-ray diffraction techniques have been used to study the structures of lipid bilayers containing basic proteins. Highly ordered multilayer specimens have been formed by using the Langmuir-Blodgett method in which a solid support is passed through a lipid monolayer held at constant surface pressure at an air/water interface. If the lipid monolayer contains acidic lipids then basic proteins in the aqueous subphase are transferred with the monolayer and incorporated into the multi-membrane stack. X-ray diffraction patterns have been recorded from multilayers of cerebroside sulphate and 40% (molar) cholesterol both with and without polylysine, cytochrome c and the basic protein from central nervous system myelin. Electron density profiles across the membranes have been derived at between 6 A and 12 A resolution. All of the membrane profiles have been placed on an absolute scale of electron density by the isomorphous exchange of cholesterol with a brominated cholesterol analog. The distributions and conformations of the various basic proteins incorporated within the cerebroside sulphate/cholesterol bilayer are very different. Polylysine attaches to the surface of the lipid bilayer as a fully extended chain while cytochrome c maintains its native structure and attaches to the bilayer surface with its short axis approximately perpendicular to the membrane plane. The myelin basic protein associates intimately with the lipid headgroups in the form of an extended molecule, yet its dimension perpendicular to the plane of the membrane of approx. 15 A is consistent with the considerable degree of secondary structure found in solution. In the membrane plane, the myelin basic protein extends to cover an area of about 2500 A2. There is no significant penetration of the protein into the hydrocarbon region of the bilayer or, indeed, beyond the position of the sulphate group of the cerebroside sulphate molecule.     

An analysis of Kohonen's self-organizing maps using a system of energy functions

In this paper a new method for analyzing Kohonen's self-organizing feature maps is presented. The method makes use of a system of energy functions, one energy function for each processing unit. It is shown that the training process is equivalent to minimizing each energy function subject to constraints. The analysis is used to prove the formation of topologically correct maps when the inherent dimensionality of the input patterns matches that of the network. The energy equations can be used to compute the steady-state weight values of the network. In addition, the analysis allows bounds on the training parameters to be determined. Finally, examples of energy landscapes are presented to graphically show the behavior of the network.     

Free energy distributions in proteins.

Protein molecules in solution have a broad distribution of enthalpy states. A good approximation to the distribution function for enthalpy states can be calculated, using the maximum-entropy method, from the moments of the distribution that, in turn, are obtained from the experimental temperature dependence of the heat capacity. In the present paper, we show that the enthalpy probability distribution can then be formulated in terms of a free energy function that gives the free energy of the protein corresponding to a particular value of the enthalpy. By the location of the minima in this function, the free energy distribution graphically indicates the most probable values of the enthalpy for the protein. We find that the behavior of the free energy functions for proteins falls somewhere between two different cases: a two-state like function with two minima, the relative levels of the two states changing with temperature; and, a single-minimum function where the position of the minimum shifts to higher enthalpy values as the temperature is increased. We show that the temperature dependence of the free energy function can be expressed in terms of a central free energy distribution for a given, fixed temperature (which is most conveniently chosen as the temperature of the maximum in the heat capacity). The nature of this central free energy function for a given protein thus yields all of the thermodynamic behavior of that protein over the temperature range of the denaturation process.     

On the energy density of helical proteins

We solve the problem of determining the energy actions whose moduli space of extremals contains the class of Lancret helices with a prescribed slope. We first see that the energy density should be linear both in the total bending and in the total twisting, such that the ratio between the weights of them is the prescribed slope. This will give an affirmative answer to the conjecture stated in Barros and Ferrández (J Math Phys 50:103529, 2009). Then, we normalize to get the best choice for the helical energy. It allows us to show that the energy, for instance of a protein chain, does not depend on the slope and is invariant under homotopic changes of the cross section which determines the cylinder where the helix is lying. In particular, the energy of a helix is not arbitrary, but it is given as natural multiples of some basic quantity of energy.     

Free energy of proton binding in proteins

In this article we use literature data on the titration of denatured ribonuclease to test the accuracy of proton-binding distributions obtained using our recent approach employing moments. We find that using only the local slope of the titration curve at a small number of points (five, for example) we can reproduce the detailed proton-binding distribution at all pH values. Our method gives the complete proton-binding polynomial for a given protein and each coefficient in this polynomial in turn yields the free energy for binding a given number of protons in all ways to the protein. Using these net free energies, we can then compute the average proton-binding free energy per proton as a function of the fraction of protons bound. We find that this function is remarkably similar for different proteins, even for proteins that exhibit quite different titration behavior. For the special case of binding to independent sites, we obtain simple relations for the first and last terms in the free energy per-proton function. For this special case we also can calculate the distribution functions giving the probability that a molecule has a given number of positive or negative charges and the joint distribution that a molecule simultaneously has a given number of positive and negative charge.     

An analysis of determinants of amino acids substitution rates in bacterial proteins

The variation of amino acid substitution rates in proteins depends on several variables. Among these, the protein's expression level, functional category, essentiality, or metabolic costs of its amino acid residues may play an important role. However, the relative importance of each variable has not yet been evaluated in comparative analyses. To this aim, we made regression analyses combining data available on these variables and on evolutionary rates, in two well-documented model bacteria, Escherichia coli and Bacillus subtilis. In both bacteria, the level of expression of the protein in the cell was by far the most important driving force constraining the amino acids substitution rate. Subsequent inclusion in the analysis of the other variables added little further information. Furthermore, when the rates of synonymous substitutions were included in the analysis of the E. coli data, only the variable expression levels remained statistically significant. The rate of nonsynonymous substitution was shown to correlate with expression levels independently of the rate of synonymous substitution. These results suggest an important direct influence of expression levels, or at least codon usage bias for translation optimization, on the rates of nonsynonymous substitutions in bacteria. They also indicate that when a control for this variable is included, essentiality plays no significant role in the rate of protein evolution in bacteria, as is the case in eukaryotes.     

An angular velocity profile in cycling derived from mechanical energy analysis

The contributions of this article are twofold. One is procedure for determining the angular velocity profile in seated cycling that maintains the total mechanical energy of both legs constant. A five-bar linkage model (thigh, shank, foot, crank and frame) of seated (fixed hip) cycling served for the derivation of the equations to compute potential and kinetic energies of the leg segments over a complete crank cycle. With experimentally collected pedal angle data as input, these equations were used to compute the total combined mechanical energy (sum of potential and kinetic energies of the segments of both legs) for constant angular velocity pedalling at 90 rpm. Total energy varied indicating the presence of internal work. Motivated by a desire to test the hypothesis that reducing internal work in cycling will reduce energy expenditure, a procedure was developed for determining the angular velocity profile that eliminated any change in total energy. Using data recorded from five subjects, this procedure was used to determine a reference profile for an average equivalent cadence of 90 rpm. The phase of this profile is such that highest and lowest angular velocities occur when the cranks are near vertical and horizontal respectively. The second contribution is the testing of the hypothesis that the reference angular velocity profile serves to effectively reduce internal work for the subjects whose data were used to develop this profile over the range of pedalling rates (80-100 rpm) naturally preferred. In this range, the internal work was decreased a minimum of 48% relative to the internal work associated with constant angular velocity pedalling. The acceptance of this hypothesis has relevance to the protocol for future experiments which explore the effect of reduced internal work on energy expenditure in cycling.     

An HPLC procedure for the analysis of proteins in lung lavage fluid

A high-performance liquid chromatography method has been developed for fractionating the protein components of the lung's extracellular lining fluid, as sampled by bronchoalveolar lavage. With this method, 10 ml (or less) of rat bronchoalveolar lavage fluid (BALF) in phosphate-buffered saline can be quantitatively analyzed rapidly and reproducibly. This volume (25% of the lavage fluid volume from one rat using a standardized lavage technique) is made 0.2% with respect to trifluoroacetic acid (TFA) and pumped through a microBondapak C18 Radial-PAK HPLC column equilibrated with H2O/0.2% TFA. Six fractions are then eluted with a series of acetonitrile gradients and isocratic steps that progress from H2O/0.2% TFA to 65% CH3 CN/0.2% TFA. Following this, 5 additional fractions are eluted with methanol. All 11 fractions are detected by monitoring the column effluents at 206 nm and can be recovered by lyophilization since all the components of the HPLC solvent system are volatile. Nine of the 11 fractions were found to contain protein. Three of the fractions contained proteins common to the blood compartment. The largest fraction of these was albumin, followed by a fraction containing immunoglobulins. Six other protein fractions appeared to be derived from the cells of the lung inasmuch as they were not detected in plasma. Two fractions contained no protein or phospholipids, whereas the most hydrophobic protein fraction did contain phospholipids. A major phospholipid fraction containing no protein eluted early in the chromatogram and was not detectable at 206 nm. This HPLC procedure offers significant utility for identifying and quantifying alterations in several BALF constituents during the development and progression of environmentally induced lung diseases as well as other pulmonary disorders.