Structural signatures of enzyme binding pockets from order-independent surface alignment: a study of metalloendopeptidase and NAD binding proteins

Detecting similarities between local binding surfaces can facilitate identification of enzyme binding sites and prediction of enzyme functions, and aid in our understanding of enzyme mechanisms. Constructing a template of local surface characteristics for a specific enzyme function or binding activity is a challenging task, as the size and shape of the binding surfaces of a biochemical function often vary. Here we introduce the concept of signature binding pockets, which captures information on preserved and varied atomic positions at multiresolution levels. For proteins with complex enzyme binding and activity, multiple signatures arise naturally in our model, forming a signature basis set that characterizes this class of proteins. Both signatures and signature basis sets can be automatically constructed by a method called SOLAR (Signature Of Local Active Regions). This method is based on a sequence-order-independent alignment of computed binding surface pockets. SOLAR also provides a structure-based multiple sequence fragment alignment to facilitate the interpretation of computed signatures. By studying a family of evolutionarily related proteins, we show that for metzincin metalloendopeptidase, which has a broad spectrum of substrate binding, signature and basis set pockets can be used to discriminate metzincins from other enzymes, to predict the subclass of metzincins functions, and to identify specific binding surfaces. Studying unrelated proteins that have evolved to bind to the same NAD cofactor, we constructed signatures of NAD binding pockets and used them to predict NAD binding proteins and to locate NAD binding pockets. By measuring preservation ratio and location variation, our method can identify residues and atoms that are important for binding affinity and specificity. In both cases, we show that signatures and signature basis set reveal significant biological insight.     

The recognition specificity of the CHD1 chromodomain with modified histone H3 peptides

Histone tail peptides comprise the flexible portion of chromatin, the substance which serves as the packaging for the eukaryotic genome. According to the histone code hypothesis, reader protein domains (chromodomains) can recognize modifications of amino acid residues within these peptides, regulating the expression of genes. We have performed simulations on models of chromodomain helicase DNA-binding protein 1 complexed with a variety of histone H3 modifications. Binding free energies for both the overall complexes and the individual residues within the protein and peptides were computed with molecular mechanics-generalized Born surface area. The simulation results agree well with experimental data and identify several chromodomain helicase DNA-binding protein 1 residues that play key roles in the interaction with each of the H3 modifications. We identified one class of protein residues that bind to H3 in all of the complexes (generally interacting hydrophobically), and a second class of residues that bind only to particular H3 modifications (generally interacting electrostatically). Additionally, we found that modifications of H3R2 and H3T3 have a dominant effect on the binding affinity; methylation of H3K4 has little effect on the interaction strength when H3R2 or H3T3 is modified. Our findings with regard to the specificity shown by the latter class of protein residues in their binding affinity to certain modifications of H3 support the histone code hypothesis.     

Interaction of the p53 DNA-binding domain with its n-terminal extension modulates the stability of the p53 tetramer

The tetrameric tumor suppressor p53 plays a pivotal role in the control of the cell cycle and provides a paradigm for an emerging class of oligomeric, multidomain proteins with structured and intrinsically disordered regions. Many of its biophysical and functional properties have been extrapolated from truncated variants, yet the exact structural and functional role of certain segments of the protein is unclear. We found from NMR and X-ray crystallography that the DNA-binding domain (DBD) of human p53, usually defined as residues 94-292, extends beyond these domain boundaries. Trp91, in the hinge region between the disordered proline-rich N-terminal domain and the DBD, folds back onto the latter and has a cation-π interaction with Arg174. These additional interactions increase the melting temperature of the DBD by up to 2 °C and inhibit aggregation of the p53 tetramer. They also modulate the dissociation of the p53 tetramer. The absence of the Trp91/Arg174 packing presumably allows nonnative DBD-DBD interactions that both nucleate aggregation and stabilize the interface. These data have important implications for studies of multidomain proteins in general, highlighting the fact that weak ordered-disordered domain interactions can modulate the properties of proteins of complex structure.     

The structure of Arabidopsis thaliana OST1 provides insights into the kinase regulation mechanism in response to osmotic stress

SnRK [SNF1 (sucrose non-fermenting-1)-related protein kinase] 2.6 [open stomata 1 (OST1)] is well characterized at molecular and physiological levels to control stomata closure in response to water-deficit stress. OST1 is a member of a family of 10 protein kinases from Arabidopsis thaliana (SnRK2) that integrates abscisic acid (ABA)-dependent and ABA-independent signals to coordinate the cell response to osmotic stress. A subgroup of protein phosphatases type 2C binds OST1 and keeps the kinase dephosphorylated and inactive. Activation of OST1 relies on the ABA-dependent inhibition of the protein phosphatases type 2C and the subsequent self-phosphorylation of the kinase. The OST1 ABA-independent activation depends on a short sequence motif that is conserved among all the members of the SnRK2 family. However, little is known about the molecular mechanism underlying this regulation. The crystallographic structure of OST1 shows that ABA-independent regulation motif stabilizes the conformation of the kinase catalytically essential α C helix, and it provides the basis of the ABA-independent regulation mechanism for the SnRK2 family of protein kinases.     

Structure-based design of a protein immunogen that displays an HIV-1 gp41 neutralizing epitope

Antibody Z13e1 is a relatively broadly neutralizing anti-human immunodeficiency virus type 1 antibody that recognizes the membrane-proximal external region (MPER) of the human immunodeficiency virus type 1 envelope glycoprotein gp41. Based on the crystal structure of an MPER epitope peptide in complex with Z13e1 Fab, we identified an unrelated protein, interleukin (IL)-22, with a surface-exposed region that is structurally homologous in its backbone to the gp41 Z13e1 epitope. By grafting the gp41 Z13e1 epitope sequence onto the structurally homologous region in IL-22, we engineered a novel protein (Z13-IL22-2) that contains the MPER epitope sequence for use as a potential immunogen and as a reagent for the detection of Z13e1-like antibodies. The Z13-IL22-2 protein binds Fab Z13e1 with a K_d of 73 nM. The crystal structure of Z13-IL22-2 in complex with Fab Z13e1 shows that the epitope region is faithfully replicated in the Fab-bound scaffold protein; however, isothermal calorimetry studies indicate that Fab binding to Z13-IL22-2 is not a lock-and-key event, leaving open the question of whether conformational changes upon binding occur in the Fab, in Z13-IL-22, or in both.     

基于效率的医院规模经济实证分析

目的 了解医院是否存在规模经济及适宜床位规模大小。方法 利用浙江省500张床及以上综合性医院数据,用主、客观结合的方式和效率回归调整模型筛选评价指标,用数据包络分析方法和Tobit回归模型进行实证分析。结果 所有决策单元的总体效率和纯技术效率、规模效率平均得分均大于0.93;36%的单元为总体有效,65%的单元为纯技术有效,43%的单元为规模有效;43%的单元处于规模报酬不变区间,42%的单元处于报酬递增区间,15%的单元处于报酬递减区间;全要素生产率平均呈下降;效率的影响因素主要有床位、药品收入比重、设备数量、医院等级、诊次床位比、业务收支比、每床卫技数、每床固定资产、所在地区千人医生数和经济水平等。结论 多数医院存在规模经济。医院床位数在800～1 300张较为合适。提高医院经营效率可采取以下措施：合理控制床位;控制药品收入比重,注重医疗技术的提高;提高管理水平,控制固定资产过度投入;提高地区卫生资源水平和经济发展水平。     

Water transport in AQP0 aquaporin: molecular dynamics studies

A double lipid bilayer structure containing opposing tetramers of AQP0 aquaporin, in contact through extracellular face loop regions, was recently modeled using an intermediate-resolution map obtained by electron crystallographic methods. The pores of these water channels were found to be critically narrow in three regions and subsequently interpreted to be those of a closed state of the channel. The subsequent determination of a high-resolution AQP0 tetramer structure by X-ray crystallographic methods yielded a pore model featuring two of the three constrictions as noted in the EM work and water molecules within the channel pore. The extracellular-side constriction region of this AQP0 structure was significantly larger than that of the EM-based model and similar to that of the highly water permeable AQP1. The X-ray-based study of AQP0 however could not ascertain if the water molecules found in the pore were the result of water entering from one or both ends of the channel, nor whether water could freely pass through all constriction points. Additionally, this X-ray-based structure could not provide an answer to the question of whether the double lipid bilayer configuration of AQP0 could functionally maintain a water impermeable state of the channel. To address these questions we conducted molecular dynamics simulations to compare the time-dependent behavior of the AQP0 and AQP1 channels within lipid bilayers. The simulations demonstrate that AQP0, in single or double lipid bilayers, is not closed to water transport and that thermal motions of critical side-chains are sufficient to facilitate the movement of water past any of its constriction regions. These motional requirements do however lead to significant free energy barriers and help explain physiological observations that found water permeability in AQP0 to be substantially lower than in the AQP1 pore.     

Recent Development of Ni/Fe‐Based Micro/Nanostructures toward Photo/Electrochemical Water Oxidation

The oxygen evolution reaction (OER), as an important process involved in water splitting and rechargeable metal–air batteries, has drawn increasing attention in the context of clean energy generation and efficient energy storage. This review concerns the progress and new discoveries in the field of Ni/Fe‐based micro/nanostructures toward electrochemical and photo‐electrochemical (PEC) water oxidation during last few years. First, toward the design and construction of new electrocatalysis, different types of current Ni/Fe‐based compounds for OER are summarized. The mechanism studies of the active phases and positions of Ni/Fe‐based micro/nanostructures are further introduced to understand the properties of catalytic active sites, which could facilitate further improving the performance of Ni/Fe‐based OER electrocatalysts. Second, splitting water using sunlight with low overpotential is another important target in making solar‐to‐hydrogen micro/nanodevices, and thus attention is then focused on the development of several important Ni/Fe‐based PEC catalysts. Third, the recent theoretical calculations on the OER mechanism during water splitting and insights into electronic structures are analyzed; finally, the future trends and perspectives are also discussed briefly.