Viral envelope glycoproteins swing into action

AbstractAnalysis of tick-borne encephalitis virus E protein reveals considerable structural diversity in the glycoproteins that clothe enveloped viruses and hints at the conformational gyrations in this molecule that lead to viral fusion

Biophysical characterization and crystal structure of the Feline Immunodeficiency Virus p15 matrix protein

International audienc

X-ray diffraction reveals the intrinsic difference in the physical properties of membrane and soluble proteins.

Membrane proteins are distinguished from soluble proteins by their insertion into biological membranes. This insertion is achieved via a noticeable arrangement of hydrophobic amino acids that are exposed at the surface of the protein, and renders the interaction with the aliphatic tails of lipids more energetically favorable. This important difference between these two categories of proteins is the source of the need for a specific handling of membrane proteins, which transpired in the creation of new tools for their recombinant expression, purification and even crystallization. Following this line, we show here that crystals of membrane proteins display systematically higher diffraction anisotropy than those of soluble proteins. This phenomenon dramatically hampers structure solution and refinement, and has a strong impact on the quality of electron-density maps. A farther search for origins of this phenomenon showed that the type of crystallization, and thus the crystal packing, has no impact on anisotropy, nor does the nature or function of the membrane protein. Membrane proteins fully embedded within the membrane display equal anisotropy compared to the ones with extra membranous domains or fusions with soluble proteins. Overall, these results overturn common beliefs and call for a specific handling of their diffraction data

Au courant computation of the PDB to audit diffraction anisotropy of soluble and membrane proteins.

This data article makes available the informed computation of the whole Protein Data Bank (PDB) to investigate diffraction anisotropy on a large scale and to perform statistics. This data has been investigated in detail in "X-ray diffraction reveals the intrinsic difference in the physical properties of membrane and soluble proteins" [1]. Diffraction anisotropy is traditionally associated with absence of contacts in-between macromolecules within the crystals in a given direction of space. There are however many case that do not follow this empirical rule. To investigate and sort out this discrepancy, we computed diffraction anisotropy for every entry of the PDB, and put them in context of relevant metrics to compare X-ray diffraction in reciprocal space to the crystal packing in real space. These metrics were either extracted from PDB files when available (resolution, space groups, cell parameters, solvent content), or calculated using standard procedures (anisotropy, crystal contacts, presence of ligands). More specifically, we separated entries to compare soluble vs membrane proteins, and further separated the later in subcategories according to their insertion in the membrane, function, or type of crystallization (Type I vs Type II crystal packing). This informed database is being made available to investigators in the raw and curated formats that can be re-used for further downstream studies. This dataset is useful to test ideas and to ascertain hypothesis based on statistical analysis

HIV-1 Pre-Integration Complexes. Structures, Functions and Drug Design

International audienc

Études fonctionnelles et structurales de la nucléoline en complexe avec des acides nucléiques structurés en G-quartet et de la pectate-lyase PelI de la bactérie phytopathogène Erwinia chrysanthemi

Nucleolin is a classical marker of cancerous cells and which interacts with nucleic acids. The purification protocol of various fragments of the protein was optimized in order to meet requirements for crystallographic studies. The protein-G-quartet structured DNA complex was subjected to a detailed characterization using biophysical methods. At the same time, crystallization trials were performed on the protein alone and in complex with nucleic acids. The structure of a DNA structured in G-quartet has been solved. E. chrysanthemi pectate lyase PelI catalyzes the degradation of pectin by a beta-elimination chemistry. The structure of the protein has been solved in its native state and in complex with a substrate, a tetragalacturonic acid. Different mutants have been produced, characterized and crystallized, leading us to a better understanding of the reaction mechanism.La nucléoline est une protéine nucléolaire servant de marqueur des cellules cancéreuses et qui interagit avec des acides nucléiques. Le protocole de purification de divers fragments de cette protéine a été optimisé afin de l adapter aux exigences d une étude cristallographique. Une analyse de l interaction de ces fragments avec diverses séquences d ADN structurées en G-quartet a été réalisée par des méthodes biophysiques. Parallèlement, des essais de cristallisation ont été menés sur ces fragments seuls et en complexe avec des acides nucléiques. La structure d un ADN structuré en G-quartet a été obtenue. La pectate lyase PelI d E. chrysanthemi catalyse la dégradation de la pectine par un mécanisme de beta-élimination. Les structures de l enzyme seule et en complexe avec un substrat, un acide tétragalacturonique, ont été déterminées. Divers mutants ont été produits, caractérisés et cristallisés, nous permettant de mieux comprendre le mécanisme réactionnelLYON1-BU.Sciences (692662101) / SudocSudocFranceF

Probing the Role of Divalent Metal Ions in a Bacterial Psychrophilic Metalloprotease: Binding Studies of an Enzyme in the Crystalline State by X-Ray Crystallography

The psychrophilic alkaline metalloprotease (PAP) produced by a Pseudomonas bacterium isolated in Antarctica belongs to the clan of metzincins, for which a zinc ion is essential for catalytic activity. Binding studies in the crystalline state have been performed by X-ray crystallography in order to improve the understanding of the role of the zinc and calcium ions bound to this protease. Cocrystallization and soaking experiments with EDTA in a concentration range from 1 to 85 mM have resulted in five three-dimensional structures with a distinct number of metal ions occupying the ion-binding sites. Evolution of the structural changes observed in the vicinity of each cation-binding site has been studied as a function of the concentration of EDTA, as well as of time, in the presence of the chelator. Among others, we have found that the catalytic zinc ion was the first ion to be chelated, ahead of a weakly bound calcium ion (Ca 700) exclusive to the psychrophilic enzyme. Upon removal of the catalytic zinc ion, the side chains of the active-site residues His-173, His-179 and Tyr-209 shifted ∼4, 1.0, and 1.6 Å, respectively. Our studies confirm and also explain the sensitivity of PAP toward moderate EDTA concentrations and propose distinct roles for the calcium ions. A new crystal form of native PAP validates our previous predictions regarding the adaptation of this enzyme to cold environments as well as the proteolytic domain calcium ion being exclusive for PAP independent of crystallization conditions