Journal Articles

  • Structural insights into FRS2α PTB domain recognition by neurotrophin receptor TrkB
    [Jun 2014]

    Abstract

    The fibroblast growth factor receptor (FGFR) substrate 2 (FRS2) family proteins function as scaffolding adapters for receptor tyrosine kinases (RTKs). The FRS2α proteins interact with RTKs through the phosphotyrosine-binding (PTB) domain and transfer signals from the activated receptors to downstream effector proteins. Here, we report the nuclear magnetic resonance structure of the FRS2α PTB domain bound to phosphorylated TrkB. The structure reveals that the FRS2α-PTB domain is comprised of two distinct but adjacent pockets for its mutually exclusive interaction with either nonphosphorylated juxtamembrane region of the FGFR, or tyrosine phosphorylated peptides TrkA and TrkB. The new structural insights suggest rational design of selective small molecules through targeting of the two conjunct pockets in the FRS2α PTB domain. Proteins 2014; 82:1534–1541. © 2014 Wiley Periodicals, Inc.

    Categories: Journal Articles
  • Identification of the bona fide DHDPS from a common plant pathogen
    [Jun 2014]

    ABSTRACT

    Agrobacterium tumefaciens is a Gram-negative soil-borne bacterium that causes Crown Gall disease in many economically important crops. The absence of a suitable chemical treatment means there is a need to discover new anti-Crown Gall agents and also characterize bona fide drug targets. One such target is dihydrodipicolinate synthase (DHDPS), a homo-tetrameric enzyme that catalyzes the committed step in the metabolic pathway yielding meso-diaminopimelate and lysine. Interestingly, there are 10 putative DHDPS genes annotated in the A. tumefaciens genome, including three whose structures have recently been determined (PDB IDs: 3B4U, 2HMC, and 2R8W). However, we show using quantitative enzyme kinetic assays that nine of the 10 dapA gene products, including 3B4U, 2HMC, and 2R8W, lack DHDPS function in vitro. A sequence alignment showed that the product of the dapA7 gene contains all of the conserved residues known to be important for DHDPS catalysis and allostery. This gene was cloned and the recombinant product expressed and purified. Our studies show that the purified enzyme (i) possesses DHDPS enzyme activity, (ii) is allosterically inhibited by lysine, and (iii) adopts the canonical homo-tetrameric structure in both solution and the crystal state. This study describes for the first time the structure, function and allostery of the bona fide DHDPS from A. tumefaciens, which offers insight into the rational design of pesticide agents for combating Crown Gall disease. Proteins 2014;. © 2014 Wiley Periodicals, Inc.

    Categories: Journal Articles
  • Molecular simulations of β-lactoglobulin complexed with fatty acids reveal the structural basis of ligand affinity to internal and possible external binding sites
    [Jun 2014]

    ABSTRACT

    The interaction of saturated fatty acids of different length (C8:0 to C18:0) with β-lactoglobulin (βLG) was investigated by molecular dynamics simulation and docking approaches. The results shows that the presence of such ligands in the hydrophobic central cavity of βLG, known as the protein calyx, determines an enhancement of atomic fluctuations compared to the unliganded form, especially for loops at the entrance of the binding site. Concerted motions are evidenced for protein regions that could favor the binding of ligands. The mechanism of anchoring of fatty acids of different length is similar for the carboxylate head-group, through electrostatic interactions with the side chains of Lys60/Lys69. The key protein residues to secure the hydrocarbon chain are Phe105/Met107, which adapt their conformation upon ligand binding. In particular, Phe105 provides an additional hydrophobic clamp only for the tail of the two fatty acids with the longest chains, palmitic and stearic acid, which are known to bind βLG with a high affinity. The search of additional external binding sites for fatty acids, distinct from the calyx, was also carried out for palmitic acid. Two external sites with a lower affinity were identified as secondary sites, one consisting in a hydrophobic cavity allowing two distinct binding modes for the fatty acid, and the other corresponding to a surface crevice close to the protein α-helix. The overall results provide a comprehensive picture of the dynamical behaviour of βLG in complex with fatty acids, and elucidate the structural basis of the binding of these physiological ligands. © Proteins 2014;. © 2014 Wiley Periodicals, Inc.

    Categories: Journal Articles
  • Increasing stability of antibody via antibody engineering: Stability engineering on an anti-hVEGF antibody
    [Jun 2014]

    Abstract

    Antibody stability is very important for expression, activity, specificity and storage. The present knowledge of antibody structure has made it possible for a computer-aided molecule design to be used to optimize and increase antibody stability. Many computational methods have been built based on knowledge or structure, however, a good integrated engineering system has yet to be developed that combines these methods. In the current study, we designed an integrated computer-aided engineering protocol, which included several successful methods. Mutants were designed considering factors that affected stability and multiwall filter screening was used to improve the design accuracy. Using this protocol, the thermo-stability of an anti-hVEGF antibody was significantly improved. Nearly 40% of the single-point mutants proved to be more stable than the parent antibody and most of the mutations could be stacked effectively. The T50 also improved about 7°C by combinational mutation of 7 sites in the light chain and 3 sites in the heavy chain. Data indicate that the protocol is an effective method for optimization of antibody structure, especially for improving thermo-stability. This protocol could also be used to enhance the stability of other antibodies. © Proteins 2014;. © 2014 Wiley Periodicals, Inc.

    Categories: Journal Articles
  • Spot quantification in two dimensional gel electrophoresis image analysis: comparison of different approaches and presentation of a novel compound fitting algorithm
    [Jun 2014]

    Background: Various computer-based methods exist for the detection and quantification of protein spots in two dimensional gel electrophoresis images. Area-based methods are commonly used for spot quantification: an area is assigned to each spot and the sum of the pixel intensities in that area, the so-called volume, is used a measure for spot signal. Other methods use the optical density, i.e. the intensity of the most intense pixel of a spot, or calculate the volume from the parameters of a fitted function. Results: In this study we compare the performance of different spot quantification methods using synthetic and real data. We propose a ready-to-use algorithm for spot detection and quantification that uses fitting of two dimensional Gaussian function curves for the extraction of data from two dimensional gel electrophoresis (2-DE) images. The algorithm implements fitting using logical compounds and is computationally efficient. The applicability of the compound fitting algorithm was evaluated for various simulated data and compared with other quantification approaches. We provide evidence that even if an incorrect bell-shaped function is used, the fitting method is superior to other approaches, especially when spots overlap. Finally, we validated the method with experimental data of urea-based 2-DE of Abeta peptides andre-analyzed published data sets. Our methods showed higher precision and accuracy than other approaches when applied to exposure time series and standard gels. Conclusion: Compound fitting as a quantification method for 2-DE spots shows several advantages over other approaches and could be combined with various spot detection methods.The algorithm was scripted in MATLAB (Mathworks) and is available as a supplemental file.
    Categories: Journal Articles
  • A Macrocyclic Chelator with Unprecedented Th4+ Affinity
    [Jun 2014]

    Journal of the American Chemical SocietyDOI: 10.1021/ja503456r
    Categories: Journal Articles
  • Electrocatalytic O2 Reduction by [Fe-Fe]-Hydrogenase Active Site Models
    [Jun 2014]

    Journal of the American Chemical SocietyDOI: 10.1021/ja5021684
    Categories: Journal Articles
  • Single Molecule FRET Analysis of the 11 Discrete Steps of a DNA Actuator
    [Jun 2014]

    Journal of the American Chemical SocietyDOI: 10.1021/ja502580t
    Categories: Journal Articles
  • A General Synthetic Approach for Ordered Mesoporous Metal Sulfides
    [Jun 2014]

    Journal of the American Chemical SocietyDOI: 10.1021/ja504407e
    Categories: Journal Articles
  • Surface Plasmon Resonance in Gold Ultrathin Nanorods and Nanowires
    [Jun 2014]

    Journal of the American Chemical SocietyDOI: 10.1021/ja503558c
    Categories: Journal Articles
  • Caution needed with model use and assumptions [Social Sciences]
    [Jun 2014]

    The article by Simonit and Perrings (1) describes development of a spatially explicit model of ecosystem service flows associated with reforestation of the Panama Canal watershed. Critical to their study are estimates of water flows, particularly during dry seasons. We have two concerns. First, we agree with Ogden and Stallard’s...
    Categories: Journal Articles
  • 104N/E of p53 is not an adaptively selected site [Biological Sciences]
    [Jun 2014]

    The theories of molecular evolution are very helpful to yield insights into the genetic mechanisms for phenotypes or physiological roles that are difficult to discern in the laboratory. Mutations of functional genes that benefit fitness may experience fixation, especially when they originate in species living in challenging environments where selection...
    Categories: Journal Articles
  • p53 codon 104 is adaptive in Tibetan mammals [Biological Sciences]
    [Jun 2014]

    In our paper, we substantiate the fact that codon 104 variation is adaptive in highland Tibet plateau mammals (1). Liu’s letter (2) claims that codon 104 is not adaptive, which is only based on searching the sequence prediction in National Center for Biotechnology Information (NCBI) without experimental evidence. Although the...
    Categories: Journal Articles
  • Dengue virus "microRNA-like" small RNA? [Biological Sciences]
    [Jun 2014]

    In PNAS, Hussain and Asgari claim they identified a “microRNA-like” 23-nucleotide RNA expressed by Dengue virus 2 (DENV2), called viral small RNA (vsRNA)-5, in infected mosquito and mammalian cells (1). vsRNA-5, which could also be detected by Northern blot or RT-PCR, was reported to repress DENV2 replication in insect Aag2...
    Categories: Journal Articles
  • Dengue virus microRNA-like small RNA [Biological Sciences]
    [Jun 2014]

    Skalsky et al. (1) indicate that viral small RNA (vsRNA)-5 (2) was not found in previous deep sequencings, is present in very low abundance, and likely is an RNA interference (RNAi) product. With advancements in next-generation sequencing, progressively deeper sequencing results are obtained. In fact, vsRNA-5 reads were present in...
    Categories: Journal Articles
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