Structural basis of hierarchical multiple substates of a protein. II: Monte Carlo simulation of native thermal fluctuations and energy minimization

Conformational fluctuations in a globular protein, bovine pancreatic trypsin inhibitor, in the time range between picoseconds and nanoseconds are studied by a Monte Carlo simulation method. Multiple energy minima are derived from sampled conformations by minimizing their energy. They are distributed in clusters in the conformational space. A hierarchical structure is observed in the simulated dynamics. In the time range between 10(-14) and 10(-10) seconds dynamics is well represented by a superposition of vibrational motions within an energy well with transitions among minima within each cluster. Transitions among clusters take place in the time range of nanoseconds or longer.     

Use of restrained molecular dynamics in water to determine three-dimensional protein structure: prediction of the three-dimensional structure of Ecballium elaterium trypsin inhibitor II

Refinement of distance geometry (DG) structures of EETI-II (Heitz et al.: Biochemistry 28:2392-2398, 1989), a member of the squash family trypsin inhibitor, have been carried out by restrained molecular dynamics (RMD) in water. The resulting models show better side chain apolar/polar surface ratio and estimated solvation free energy than structures refined "in vacuo." The consistent lower values of residual NMR constraint violations, apolar/polar surface ratio, and solvation free energy for one of these refined structures allowed prediction of the 3D folding and disulfide connectivity of EETI-II. Except for the few first residues for which no NMR constraints were available, this computer model fully agreed with X-ray structures of CMTI-I (Bode et al.: FEBS Lett. 242:285-292, 1989) and EETI-II complexed with trypsin that appeared after the RMD simulation was completed. Restrained molecular dynamics in water is thus proved to be highly valuable for refinement of DG structures. Also, the successful use of apolar/polar surface ratio and of solvation free energy reinforce the analysis of Novotny et al. (Proteins 4:19-30, 1988) and shows that these criteria are useful indicators of correct versus misfolded models.     

Milling energy requirement of the aneuploid stocks of common wheat,including alien addition lines

Summary Aneuploid stocks, which included Triticum aestivum/alien, disomic, chromosome addition lines, wheat/alien, ditelosomic, chromosome addition lines, and the available aneuploids of Chinese Spring wheat, were used to locate genes that influence milling energy requirement (ME). Genes that affected ME were found on all seven homoeologous chromosome groups. The addition of complete wheat chromosomes 1B, 1D, 2A, 2D, 5B, 6B, 7B and 7D increased ME. Positive effects were also found in specific chromosome arms: 1BS, 2DS, 5AS, 5BS and 6BL. Wheat chromosome 3B conditioned low ME and the gene(s) responsible was located on the short arm. Other negative effects were attributed to wheat chromosome arms 4BL, 4DL, 5DS and 6DS. Alien chromosome additions that conferred high ME included 2H, 5H, 6H and 7H of barley, Hordeum vulgare and 2R, 2R, 4R, 4RL, 6R, 6RL and 7RL of rye, Secale cereale. Those that conferred a low ME included 1H ^ch of H. chilense, and 6u and 7u of Aegilops umbellulata, 5R and 5RS of S. cereale and 5R ^m and 5R ^mS of S. montanum. Although the control of ME is polygenic, there is a major effect of genes located on the short arms of homoeologous group 5 chromosomes.     

The meaning of interaction parameters in two-state protein complexes

The theory of interaction parameters has thus far been based on the free-energy relationships in the formation of ternary complexes formed between a pair of ligands and a protein molecule. The concept has been formulted in terms of a thermodynamic square comprised of the free protein, the two binary complexes, and the ternary complex. However, an increasing number of proteins have been found to exist as equilibrium mixtures of two macrostates. The equilibrium constants for such two-state transitions vary quite considerably between the various binary and ternary complexes of a given protein. We show here that the interpretations of interaction parameters in such two-state systems, requiring the use of a thermodynamic cube, are much more complex than those based on the classic thermodynamic square commonly employed. We demonstrate the use of enthalpies of interaction and heat capacities of interaction to analyze the source of observed free enerigies of interaction in such systems. Specifically, we find that measured negative interaction parameters may arise simply from the inability of a system to achieve all of the positive component effects anticipated by the conventional formulation.     

On the nature of the photosynthetic energy storage monitored by photoacoustic spectroscopy

The photosynthetic energy storage yield of uncoupled thylakoid membranes was monitored by photoacoustic spectroscopy at various measuring beam intensities. The energy storage rate as evaluated by the half-saturation measuring beam intensity (i₅₀) was inhibited by 3-(3,4-dichlorophenyl)-1,1 dimethylurea, by heat inactivation or by artificial electron acceptors specific for photosystem I or photosystem II; and was activated by electron donors to photosystem I. The reactions involving both photosystems were all characterized by a similar maximal energy storage yield of 16±2 percent. The data could be interpreted if we assumed that the energy storage elicited by the photosystems at 35 Hz is detected at the level of the plastoquinone pool.Abbreviations PS photosystem - Tes N-Tris [hydroxymethl] methyl-2-aminoethanesulfonic acid - DCMU 3-(3,4-dichlorophenyl)-1,1-dimethylurea - DCIP 2,6-dichlorophenolindophenol - FeCN potassium ferricyanide - DCBQ 2,5-dichlorobenzoquinone - TMPD N,N,N-tetramethyl-p-phenilenediamine     

湘南山区农业生态系统能量分析

一、引言由于农业系统的复杂性和存在较多的随机因素,对农业系统实行全面定量研究是比较困难的,仅从定性的描述和一些经济指标来判定系统的优劣及系统的特征是不理想的。而从能量观点对农业生态系统加以分析则是一个全面且能定量的方法,并且具有较强的客观性,能够反映农业生态系统最基本、最本质的关系。     

Calcium binding to the subunit c ofE. coli ATP-synthase and possible functional implications in energy coupling

The 8-kDa subunit c of theE. coli F₀ ATP-synthase proton channel was tested for Ca⁺⁺ binding activity using a⁴⁵Ca⁺⁺ ligand blot assay after transferring the protein from SDS-PAGE gels onto polyvinyl difluoride membranes. The purified subunit c binds⁴⁵Ca⁺⁺ strongly with Ca⁺⁺ binding properties very similar to those of the 8-kDa CF₀ subunit III of choloroplast thylakoid membranes. The N-terminal f-Met carbonyl group seems necessary for Ca⁺⁺ binding capacity, shown by loss of Ca⁺⁺ binding following removal of the formyl group by mild acid treatment. The dicyclohexylcarbodiimide-reactive Asp-61 is not involved in the Ca⁺⁺ binding, shown by Ca⁺⁺ binding being retained in twoE. coli mutants, Asp61Asn and Asp61Gly. The Ca⁺⁺ binding is pH dependent in both theE. coli and thylakoid 8-kDa proteins, being absent at pH 5.0 and rising to a maximum near pH 9.0. A treatment predicted to increase the Ca⁺⁺ binding affinity to its F₀ binding site (chlorpromazine photoaffinity attachment) caused an inhibition of ATP formation driven by a base-to-acid pH jump in whole cells. Inhibition was not observed when the Ca⁺⁺ chelator EGTA was present with the cells during the chlorpromazine photoaffinity treatment. An apparent Ca⁺⁺ binding constant on the site responsible for the UV plus chlorpromazine effect of near 80–100 nM was obtained using an EGTA-Ca⁺⁺ buffer system to control free Ca⁺⁺ concentration during the UV plus chlorpromazine treatment. The data are consistent with the notion that Ca⁺⁺ bound to the periplasimic side of theE. coli F₀ proton channel can block H⁺ entry into the channel. A similar effect occurs in thylakoid membranes, but the Ca⁺⁺ binding site is on the lumen side of the thylakoid, where Ca⁺⁺ binding can modulate acid-base jump ATP formation. The Ca⁺⁺ binding to the F₀ and CF₀ complexes is consistent with a pH-dependent gating mechanism for control of H⁺ ion flux across the opening of the H⁺ channel.This work was supported in part by grants from the Department of Energy and the U.S. Department of Agriculture.On leave from the Institute of Soil Science and Photosynthesis, Russian Academy of Science, Pushchino, Russia.     

Control of Photosystem II in spinach leaves by continuous light and by light pulses given in the dark

The light-induced induction of components of non-photochemical quenching of chlorophyll fluorescence which are distinguished by different rates of dark relaxation (qNf, rapidly relaxing and qNs, slowly relaxing or not relaxing at all in the presence brief saturating light pulses which interrupt darkness at low frequencies) was studied in leaves of spinach.After dark adaptation of the leaves, a fast relaxing component developed in low light only after a lag phase. Quenching increased towards a maximum with increasing photon flux density. This fast component of quenching was identified as energy-dependent quenching qE. It required formation of an appreciable transthylakoid pH and was insignificant when darkened spinach leaves received 1 s pulses of light every 30 s even though zeaxanthin was formed from violaxanthin under these conditions.Another quenching component termed qNs developed in low light without a lag phase. It was not dependent on a transthylakoid pH gradient, decayed exponentially with a long half time of relaxation and was about 20% of total quenching irrespective of light intensity. When darkened leaves were flashed at frequencies higher than 0.004 Hz with 1 s light pulses, this quenching also appeared. Its extent was very considerable, and it did not require formation of zeaxanthin. Relaxation was accelerated by far-red light, and this acceleration was abolished by NaF.We suggest that qNs is the result of a so-called state transition, in which LHC II moves after its phosphorylation from fluorescent PS II to nonfluorescent PS I. This state transition was capable of decreasing in darkened leaves the potential maximum quantum efficiency of electron flow through Photosystem II by about 20%.Abbreviations PFD photon flux density - PS photosystem     

Modeling the temperature response of nitrification

To model nitrification rates in soils, it is necessary to have equations that accurately describe the effect of environmental variables on nitrification rates. A variety of equations have been used previously to describe the effect of temperature on rates of microbial processes. It is not clear which of these best describes the influence of temperature on nitrification rates in soil. I compared five equations for describing the effects of temperature on nitrification in two soils with very different temperature optima from a California oak woodland-annual grassland. The most appropriate equation depended on the range of temperatures being evaluated. A generalized Poisson density function best described temperature effects on nitrification rates in both soils over the range of 5 to 50 °C; however, the Arrhenius equation best described temperature effects over the narrower range of soil temperatures that normally occurs in the ecosystem (5 to 28 °C). Temperature optima for nitrification in most of the soils were greater than even the highest soil temperatures recorded at the sites. A model accounting for increased maintenance energy requirements at higher temperatures demonstrates how net energy production, rather than the gross energy production from nitrification, is maximized during adaptation by nitrifier populations to soil temperatures.