Exploring the thermal stability and activity of α-chymotrypsin in the presence of spermine

Polyamines such as spermine can have interaction with protein. The aim of the present study was to investigate how spermine could influence the structure, thermal stability, and the activity of α-chymotrypsin. Kinetics, thermodynamics, molecular dynamics (MD), and docking simulations studies were conducted to investigate the effect of spermine on the activity and structure of α-Chymotrypsin (α-Chy) in 50 mM Tris–HCl buffer, with the pH 8, using different spectroscopic techniques as well as molecular docking and MD simulations. The stability and activity of α-Chy were increased in the presence of spermine. The results of the kinetic study showed that the activity of spermine was increased. Enzyme activation was accompanied by changes on the α-Chy conformation. Fluorescence intensity changes showed dynamic quenching during spermine binding. The fluorescence quenching of the α-Chy suggested the more polar location of Trp residues. Near-UV and Far-UV circular dichroism studies also demonstrated the transfer of Trp, Phe, and Tyr residues to a more flexible environment. The increase in the absorption of α-Chy in the presence of spermine was as a result of the formation of spermine–α-Chy complex. Molecular docking results revealed the presence of one binding site with a negative value for the Gibbs free energy of the binding of spermine to α-Chy. Docking study also revealed that van der Waals interactions and hydrogen bonds played a major role in stabilizing the complex.     

Multiple molecular dynamics simulations of human LOX‐1 and Trp150Ala mutant reveal the structural determinants causing the full deactivation of the receptor

Multiple classical molecular dynamics simulations have been applied to the human LOX‐1 receptor to clarify the role of the Trp150Ala mutation in the loss of binding activity. Results indicate that the substitution of this crucial residue, located at the dimer interface, markedly disrupts the wild‐type receptor dynamics. The mutation causes an irreversible rearrangement of the subunits interaction pattern that in the wild‐type protein allows the maintaining of a specific symmetrical motion of the monomers. The subunits dislocation determines a loss of linearity of the arginines residues composing the basic spine and a consequent alteration of the long‐range electrostatic attraction of the substrate. Moreover, the anomalous subunits arrangement observed in the mutated receptor also affects the integrity of the hydrophobic tunnel, actively involved in the short‐range hydrophobic recognition of the substrate. The combined effect of these structural rearrangements generates the impairing of the receptor function.     

Altered dynamics upon oligomerization corresponds to key functional sites

It is known that over half of the proteins encoded by most organisms function as oligomeric complexes. Oligomerization confers structural stability and dynamics changes in proteins. We investigate the effects of oligomerization on protein dynamics and its functional significance for a set of 145 multimeric proteins. Using coarse‐grained elastic network models, we inspect the changes in residue fluctuations upon oligomerization and then compare with residue conservation scores to identify the functional significance of these changes. Our study reveals conservation of about ½ of the fluctuations, with ¼ of the residues increasing in their mobilities and ¼ having reduced fluctuations. The residues with dampened fluctuations are evolutionarily more conserved and can serve as orthosteric binding sites, indicating their importance. We also use triosephosphate isomerase as a test case to understand why certain enzymes function only in their oligomeric forms despite the monomer including all required catalytic residues. To this end, we compare the residue communities (groups of residues which are highly correlated in their fluctuations) in the monomeric and dimeric forms of the enzyme. We observe significant changes to the dynamical community architecture of the catalytic core of this enzyme. This relates to its functional mechanism and is seen only in the oligomeric form of the protein, answering why proteins are oligomeric structures. Proteins 2017; 85:1422–1434. © 2017 Wiley Periodicals, Inc.     

Fast further purification of diastereomeric salts of a nonracemic acid by gas antisolvent fractionation

A novel, green possibility of the further purification of the diastereomeric salt of 4‐chloromandelic acid and 1‐phenylethane‐1‐amine was developed. Gas antisolvent method using supercritical carbon dioxide was applied for the first time to precipitate the diastereomeric salts with increased purity followed by the supercritical fluid extraction of the dissolved diastereomers. The RR ‐salt can be purified to >99%, while fractionation‐based purification of the SR ‐salt is limited to ~80%. The limiting initial diastereomeric excess correlates strongly with the atmospheric melting eutectic composition of the same salts, which suggests that despite the fast precipitation, the diastereomeric excess of the solid product is not kinetically determined. The efficiency of the diastereomeric enrichment is in the same range as that of the atmospheric reference experiments; however, technological advantages provided by the antisolvent precipitation method such as fast processing and dry product obtained suggest that this novel procedure is a promising alternative to the atmospheric methods.     

采用稳定碳同位素法分析白羊草在不同干旱胁迫下的水分利用效率

本研究以黄土高原乡土草种白羊草(Bothriochloa ischaemum(L.)Keng.)为研究对象,采用盆栽控制实验,比较白羊草在3个水分处理(CK80%FC、MS60%FC和SS 40%FC)下的生物量积累和分配模式、瞬时水分利用效率(WUE)、不同部位(新叶、老叶、茎、细根、粗根)的稳定碳同位素组成(δ~(13)C)和碳同位素分辨率(Δ~(13)C)及其相互关系,以及干旱胁迫下影响水分利用效率的主导环境因子。结果表明:1)重度干旱胁迫显著降低植物整体生物量,显著增加根冠比和细根生物量比例;2)随着干旱胁迫加剧,白羊草各器官的δ~(13)C均呈上升趋势,Δ~(13)C呈减小趋势,SS处理不同器官δ~(13)C和Δ~(13)C没有显著差异,CK和MS处理的各器官δ~(13)C均值表现分别为细根粗根老叶新叶茎、细根新叶老叶粗根茎,CK和MS处理Δ~(13)C的值总体呈根叶茎。3)新叶的δ~(13)CNL和Δ~(13)CNL与WUE的相关系数均最大,说明利用稳定碳同位素方法测定白羊草水分利用效率具有可行性。4)不同水分处理的WUE的主导影响因子不同,CK、MS、SS水分处理WUE分别受到叶面温度、大气水汽压亏缺和空气温度的影响最大。为采用稳定碳同位素方法指示白羊草水分利用效率可行性及阐明植物的胁迫响应机制提供理论依据。     

Ultrafine Metal Nanoparticles/N‐Doped Porous Carbon Hybrids Coated on Carbon Fibers as Flexible and Binder‐Free Water Splitting Catalysts

By employing in situ reduction of metal precursor and metal‐assisted carbon etching process, this study achieves a series of ultrafine transition metal‐based nanoparticles (Ni–Fe, Ni–Mo) embedded in N‐doped carbon, which are found efficient catalysts for electrolytic water splitting. The as‐prepared hybrid materials demonstrate outstanding catalytic activities as non‐noble metal electrodes rendered by the synergistic effect of bimetal elements and N‐dopants, the improved electrical conductivity, and hydrophilism. Ni/Mo₂C@N‐doped porous carbon (NiMo‐polyvinylpyrrolidone (PVP)) and NiFe@N‐doped carbon (NiFe‐PVP) produce low overpotentials of 130 and 297 mV at a current density of 10 mA cm^?2 as catalysts for hydrogen evolution reaction and oxygen evolution reaction, respectively. In addition, these binder‐free electrodes show long‐term stability. Overall water splitting is also demonstrated based on the couple of NiMo‐PVP||NiFe‐PVP catalyzer. This represents a simple and effective synthesis method toward a new type of nanometal–carbon hybrid electrodes.     

The dynamic properties of the Hepatitis C Virus E2 envelope protein unraveled by molecular dynamics

Hepatitis C Virus (HCV) is one of the most persistent human viruses. Although effective therapeutic approaches have been recently discovered, their use is limited by the elevated costs. Therefore, the development of alternative/complementary strategies is an urgent need. The E2 glycoprotein, the most immunogenic HCV protein, and its variants represent natural candidates to achieve this goal. Here we report an extensive molecular dynamics (MD) analysis of the intrinsic properties of E2. Our data provide interesting clues on the global and local intrinsic dynamic features of the protein. Present MD data clearly indicate that E2 combines a flexible structure with a network of covalent bonds. Moreover, the analysis of the two most important antigenic regions of the protein provides some interesting insights into their intrinsic structural and dynamic properties. Our data indicate that a fluctuating β-hairpin represents a populated state by the region E2^412?423. Interestingly, the analysis of the epitope E2^427?446 conformation, that undergoes a remarkable rearrangement in the simulation, has significant similarities with the structure that the E2^430?442 fragment adopts in complex with a neutralizing antibody. Present data also suggest that the strict conservation of Gly436 in E2 protein of different HCV genotypes is likely dictated by structural restraints. Moreover, the analysis of the E2^412?423 flexibility provides insights into the mechanisms that some antibodies adopt to anchor Trp437 that is fully buried in E2. Finally, the present investigation suggests that MD simulations should systematically complement crystallographic studies on flexible proteins that are studied in combination with antibodies.     

Loss of CpSRP54 function leads to a truncated light-harvesting antenna size in Chlamydomonas reinhardtii

The Chlamydomonas reinhardtii truncated light-harvesting antenna 4 (tla4) DNA transposon mutant has a pale green phenotype, a lower chlorophyll (Chl) per cell and a higher Chl a/b ratio in comparison with the wild type. It required a higher light intensity for the saturation of photosynthesis and displayed a greater per chlorophyll light-saturated rate of oxygen evolution than the wild type. The Chl antenna size of the photosystems in the tla4 mutant was only about 65% of that measured in the wild type. Molecular genetic analysis revealed that a single plasmid DNA insertion disrupted two genes on chromosome 11 of the mutant. A complementation study identified the “chloroplast signal recognition particle 54” gene (CpSRP54), as the lesion causing the tla4 phenotype. Disruption of this gene resulted in partial failure to assemble and, therefore, lower levels of light-harvesting Chl-binding proteins in the C. reinhardtii thylakoids. A comparative in silico 3-D structure-modeling analysis revealed that the M-domain of the CpSRP54 of C. reinhardtii possesses a more extended finger loop structure, due to different amino acid composition, as compared to that of the Arabidopsis CpSRP54. The work demonstrated that CpSRP54 deletion in microalgae can serve to generate tla mutants with a markedly smaller photosystem Chl antenna size, improved solar energy conversion efficiency, and photosynthetic productivity in high-density cultures under bright sunlight conditions.     

广西不同林龄杉木、马尾松人工林根系生物量及碳储量特征

为了解不同林龄杉木、马尾松人工林地地下根系生物量及碳储量特征,以广西杉木、马尾松主产区5个不同林龄阶段(幼龄林、中龄林、近熟林、成熟林、过熟林)的人工林为研究对象,采用全根挖掘法和土钻法获取标准木根系生物量、灌草根系生物量和林分细根生物量,并测定其碳含量,分析其不同林龄阶段地下根系生物量和碳储量分配特征。结果表明:杉木、马尾松林地下根系总生物量分别在9.06—31.40Mg/hm~2和7.91—53.40Mg/hm~2之间,各林龄阶段根系总生物量总体上呈现随林龄增加而增加的趋势,杉木林细根生物量随林龄的增加呈现出先减后增的趋势,马尾松呈现出逐渐减小的趋势;林分各层次根系碳含量表现为乔木灌木草本、细根;杉木、马尾松地下根系碳储量变化趋势与生物量变化趋势相同,杉木、马尾松林不同林龄阶段各层次根系和土壤细根总碳储量分别在7.56—21.97Mg/hm~2和8.86—29.95Mg/hm~2之间;地下根系碳储量总体上以乔木根系占优势,且随林龄的增大其比例呈增加的趋势。