scholarly journals Combining random microseed matrix screening and the magic triangle for the efficient structure solution of a potential lysin from bacteriophage P68

2019 ◽  
Vol 75 (7) ◽  
pp. 670-681
Author(s):  
Jia Quyen Truong ◽  
Santosh Panjikar ◽  
Linda Shearwin-Whyatt ◽  
John B. Bruning ◽  
Keith E. Shearwin
Keyword(s):  
Hen Egg White Lysozyme ◽  
High Quality ◽  
X Ray Crystallography ◽  
Egg White Lysozyme ◽  
Structure Solution ◽  
Experimental Phasing ◽  
Sad Phasing ◽  
Hen Egg White ◽  
Novel Structures

Two commonly encountered bottlenecks in the structure determination of a protein by X-ray crystallography are screening for conditions that give high-quality crystals and, in the case of novel structures, finding derivatization conditions for experimental phasing. In this study, the phasing molecule 5-amino-2,4,6-triiodoisophthalic acid (I3C) was added to a random microseed matrix screen to generate high-quality crystals derivatized with I3C in a single optimization experiment. I3C, often referred to as the magic triangle, contains an aromatic ring scaffold with three bound I atoms. This approach was applied to efficiently phase the structures of hen egg-white lysozyme and the N-terminal domain of the Orf11 protein from Staphylococcus phage P68 (Orf11 NTD) using SAD phasing. The structure of Orf11 NTD suggests that it may play a role as a virion-associated lysin or endolysin.

scholarly journals Structure Determination of Hen Egg-White Lysozyme Aggregates Adsorbed to Lipid/Water and Air/Water Interfaces

Langmuir ◽  
2020 ◽  
Vol 36 (17) ◽  
pp. 4766-4775
Author(s):  
S. Strazdaite ◽  
E. Navakauskas ◽  
J. Kirschner ◽  
T. Sneideris ◽  
G. Niaura
Keyword(s):  
Structure Determination ◽  
Egg White ◽  
Hen Egg White Lysozyme ◽  
Egg White Lysozyme ◽  
Hen Egg White ◽  
Hen Egg

Simplified heavy-atom derivatization of protein structures via co-crystallization with the MAD tetragon tetrabromoterephthalic acid

2021 ◽  
Vol 77 (5) ◽  
pp. 156-162
Author(s):  
Jia Q. Truong ◽  
Stephanie Nguyen ◽  
John B. Bruning ◽  
Keith E. Shearwin
Keyword(s):  
Heavy Atom ◽  
Protein Structures ◽  
Molecular Replacement ◽  
Hen Egg White Lysozyme ◽  
Phase Problem ◽  
Egg White Lysozyme ◽  
Mad Phasing ◽  
Experimental Phasing ◽  
Anomalous Signal ◽  
Hen Egg White

The phase problem is a persistent bottleneck that impedes the structure-determination pipeline and must be solved to obtain atomic resolution crystal structures of macromolecules. Although molecular replacement has become the predominant method of solving the phase problem, many scenarios still exist in which experimental phasing is needed. Here, a proof-of-concept study is presented that shows the efficacy of using tetrabromoterephthalic acid (B4C) as an experimental phasing compound. Incorporating B4C into the crystal lattice using co-crystallization, the crystal structure of hen egg-white lysozyme was solved using MAD phasing. The strong anomalous signal generated by its four Br atoms coupled with its compatibility with commonly used crystallization reagents render B4C an effective experimental phasing compound that can be used to overcome the phase problem.

The metalation of hen egg white lysozyme impacts protein stability as shown by ion mobility mass spectrometry, differential scanning calorimetry, and X-ray crystallography

2017 ◽  
Vol 53 (30) ◽  
pp. 4246-4249 ◽  
Author(s):  
Matthew P. Sullivan ◽  
Michael Groessl ◽  
Samuel M. Meier ◽  
Richard L. Kingston ◽  
David C. Goldstone ◽  
...  
Keyword(s):  
Ion Mobility ◽  
Egg White ◽  
Ion Mobility Mass Spectrometry ◽  
Hen Egg White Lysozyme ◽  
Scanning Calorimetry ◽  
X Ray ◽  
X Ray Crystallography ◽  
Egg White Lysozyme ◽  
Hen Egg White ◽  
Hen Egg

Metalation of lysozyme with anticancer organometallics results in protein destabilisation, probably relevant in metallodrug mode of action.

Isothermal compressibility of macromolecular crystals and macromolecules derived from high-pressure X-ray crystallography

2010 ◽  
Vol 43 (3) ◽  
pp. 407-416 ◽  
Author(s):  
Isabella Ascone ◽  
Richard Kahn ◽  
Eric Girard ◽  
Thierry Prangé ◽  
Anne-Claire Dhaussy ◽  
...  
Keyword(s):  
High Pressure ◽  
Crystal Structures ◽  
Cell Mass ◽  
Complete Information ◽  
Unit Cell ◽  
Cell Content ◽  
X Ray Crystallography ◽  
Egg White Lysozyme ◽  
Hen Egg White

The compressibility of several nucleic acid and globular protein crystals has been investigated by high-pressure macromolecular crystallography. Further, crystal structures at four different pressures allowed the determination of the intrinsic compressibilityversuspressure of d(GGTATACC)2and hen egg-white lysozyme. For lysozyme, the values for the intrinsic molecular compressibility at atmospheric pressure and the nonlinearity index were 0.070 GPa−1and 8.15, respectively. On the basis of two crystal structures at atmospheric and high pressure, similar, albeit less complete, information was derived for d(CGCGAATTCGCG)2and bovine erythrocyte Cu,Zn superoxide dismutase. Using these data and accurate calculations of the solvent-excluded volume, the apparent solvent compressibility in the crystalline state was determined as a function of pressure and compared with results from a simple model that assumes invariant unit-cell content, with the conclusion that solvent compressibility was abnormal for three out of the five crystals investigated. Experimental results suggest that macromolecular crystals submitted to high pressure may have a variable unit-cell mass due to solvent exchange with the surrounding pool, as already observed in other hydrated crystals such as zeolites.

scholarly journals Proline/alanine-rich sequence (PAS) polypeptides as an alternative to PEG precipitants for protein crystallization

2020 ◽  
Vol 76 (7) ◽  
pp. 320-325 ◽  
Author(s):  
André Schiefner ◽  
Rebecca Walser ◽  
Michaela Gebauer ◽  
Arne Skerra
Keyword(s):  
Tertiary Structure ◽  
Protein Crystallization ◽  
Repetitive Sequences ◽  
Hen Egg White Lysozyme ◽  
X Ray Crystallography ◽  
Egg White Lysozyme ◽  
Hen Egg White ◽  
Biomolecular Crystallography ◽  
Macromolecular Composition ◽  
High Diffraction

Proline/alanine-rich sequence (PAS) polypeptides represent a novel class of biosynthetic polymers comprising repetitive sequences of the small proteinogenic amino acids L-proline, L-alanine and/or L-serine. PAS polymers are strongly hydrophilic and highly soluble in water, where they exhibit a natively disordered conformation without any detectable secondary or tertiary structure, similar to polyethylene glycol (PEG), which constitutes the most widely applied precipitant for protein crystallization to date. To investigate the potential of PAS polymers for structural studies by X-ray crystallography, two proteins that were successfully crystallized using PEG in the past, hen egg-white lysozyme and the Fragaria × ananassa O-methyltransferase, were subjected to crystallization screens with a 200-residue PAS polypeptide. The PAS polymer was applied as a precipitant using a vapor-diffusion setup that allowed individual optimization of the precipitant concentration in the droplet in the reservoir. As a result, crystals of both proteins showing high diffraction quality were obtained using the PAS precipitant. The genetic definition and precise macromolecular composition of PAS polymers, both in sequence and in length, distinguish them from all natural and synthetic polymers that have been utilized for protein crystallization so far, including PEG, and facilitate their adaptation for future applications. Thus, PAS polymers offer potential as novel precipitants for biomolecular crystallography.

Neutron protein crystallography at ultra-low (<15 K) temperatures

2012 ◽  
Vol 45 (4) ◽  
pp. 686-692 ◽  
Author(s):  
Dean A. A. Myles ◽  
François Dauvergne ◽  
Matthew P. Blakeley ◽  
Flora Meilleur
Keyword(s):  
Neutron Diffraction ◽  
Protein Crystallography ◽  
Egg White ◽  
Hen Egg White Lysozyme ◽  
Large Protein ◽  
High Quality ◽  
Egg White Lysozyme ◽  
Hen Egg White ◽  
Neutron Protein Crystallography ◽  
Scientific Questions

Techniques and equipment have been developed that enable large protein crystals (1–6 mm3) flash-cooled in liquid nitrogen at 77 K to be transferred and mounted on a liquid helium Displex cryorefrigerator and cooled to temperatures down to 15 K for accurate neutron diffraction analysis. In preliminary experiments, it was possible to collect high-quality high-resolution neutron diffraction data to 1.55 Å resolution from several large crystals of triclinic hen egg white lysozyme cooled to 15 K. This enabled the subsequent cryogenic analysis of two further proteins, rubredoxin and concanavalin A, at 1.7 and 2.5 Å, respectively, demonstrating the generality of the approach. The ability to flash-cool such large crystals for cryogenic neutron analysis should significantly broaden the range of scientific questions examined by neutron protein crystallography, allowing the analysis of structures and transitions as a function of temperature and enabling freeze-trapped capture of kinetic intermediates in protein systems.

An improved method for protein crystal density measurements

1999 ◽  
Vol 32 (5) ◽  
pp. 1006-1009 ◽  
Author(s):  
Adelaine K. W. Leung ◽  
Michael M. V. Park ◽  
David W. Borhani
Keyword(s):  
Protein Crystal ◽  
Protein Crystals ◽  
Density Gradients ◽  
Hen Egg White Lysozyme ◽  
Human Apolipoprotein ◽  
Egg White Lysozyme ◽  
Crystal Density ◽  
Solvent Density ◽  
Hen Egg White

Determination of the density of protein crystals by flotation in organic solvent density gradients using simple methods for the preparation of the density gradients and the transfer of crystals into these gradients is described. The method was tested with crystals of hen egg white lysozyme. These methods are especially suitable for use with fragile, high-solvent-content protein crystals. These methods were applied to the measurement of the density of human apolipoprotein A-I crystals.

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