Using graphlet degree vectors to predict atomic displacement parameters in protein structures
In structural biology, atomic displacement parameters, commonly used in the form of B values, describe uncertainties in atomic positions. Their distribution over the structure can provide hints on local structural reliability and mobility. A spatial macromolecular model can be represented by a graph whose nodes are atoms and whose edges correspond to all interatomic contacts within a certain distance. Small connected subgraphs, called graphlets, provide information about the wiring of a particular atom. The multiple linear regression approach based on this information aims to predict a distribution of values of isotropic a...
Source: Acta Crystallographica Section D - November 21, 2023 Category: Biochemistry Authors: Pra ž nikar, J. Tags: atomic displacement parameters graphlet degree vectors interatomic contacts macromolecules research papers Source Type: research
Structural mechanism of Escherichia coli cyanase
Cyanase plays a vital role in the detoxification of cyanate and supplies a continuous nitrogen source for soil microbes by converting cyanate to ammonia and carbon dioxide in a bicarbonate-dependent reaction. The structures of cyanase complexed with dianion inhibitors, in conjunction with biochemical studies, suggest putative binding sites for substrates. However, the substrate-recognition and reaction mechanisms of cyanase remain unclear. Here, crystal structures of cyanase from Escherichia coli were determined in the native form and in complexes with cyanate, bicarbonate and intermediates at 1.5 – 1.9 Å resolution...
Source: Acta Crystallographica Section D - November 16, 2023 Category: Biochemistry Authors: Kim, J. Kim, Y. Park, J. Nam, K.H. Cho, Y. Tags: cyanase Escherichia coli substrate recognition X-ray free-electron lasers intermediate structures gating mechanisms research papers Source Type: research
Improved joint X-ray and neutron refinement procedure in Phenix
Neutron diffraction is one of the three crystallographic techniques (X-ray, neutron and electron diffraction) used to determine the atomic structures of molecules. Its particular strengths derive from the fact that H (and D) atoms are strong neutron scatterers, meaning that their positions, and thus protonation states, can be derived from crystallographic maps. However, because of technical limitations and experimental obstacles, the quality of neutron diffraction data is typically much poorer (completeness, resolution and signal to noise) than that of X-ray diffraction data for the same sample. Further, refinement is more...
Source: Acta Crystallographica Section D - November 9, 2023 Category: Biochemistry Authors: Liebschner, D. Afonine, P.V. Poon, B.K. Moriarty, N.W. Adams, P.D. Tags: macromolecular crystallography neutron diffraction joint XN refinement research papers Source Type: research
The bad and the good of trends in model building and refinement for sparse-data regions: pernicious forms of overfitting versus good new tools and predictions
Model building and refinement, and the validation of their correctness, are very effective and reliable at local resolutions better than about 2.5 Å for both crystallography and cryo-EM. However, at local resolutions worse than 2.5 Å both the procedures and their validation break down and do not ensure reliably correct models. This is because in the broad density at lower resolution, critical features such as protein backbone carbonyl O atoms are not just less accurate but are not seen at all, and so peptide orientations are frequently wrongly fitted by 90 – 180 ° . This puts both backbone and side chains into...
Source: Acta Crystallographica Section D - November 3, 2023 Category: Biochemistry Authors: Richardson, J.S. Williams, C.J. Chen, V.B. Prisant, M.G. Richardson, D.C. Tags: overfitting at low resolution machine-learning predictions CaBLAM validation ISOLDE rebuilding Ramachandran refinement problems research papers Source Type: research
Neutron crystallographic refinement with REFMAC5 from the CCP4 suite
Hydrogen (H) atoms are abundant in macromolecules and often play critical roles in enzyme catalysis, ligand-recognition processes and protein – protein interactions. However, their direct visualization by diffraction techniques is challenging. Macromolecular X-ray crystallography affords the localization of only the most ordered H atoms at (sub-)atomic resolution (around 1.2 Å or higher). However, many H atoms of biochemical significance remain undetectable by this method. In contrast, neutron diffraction methods enable the visualization of most H atoms, typically in the form of deuterium (2H) atoms, at much more co...
Source: Acta Crystallographica Section D - November 3, 2023 Category: Biochemistry Authors: Catapano, L. Long, F. Yamashita, K. Nicholls, R.A. Steiner, R.A. Murshudov, G.N. Tags: neutron macromolecular crystallography crystallographic refinement H atoms REFMAC5 CCP4 research papers Source Type: research
Cocrystallization of ubiquitin – deubiquitinase complexes through disulfide linkage
Structural characterization of the recognition of ubiquitin (Ub) by deubiquitinases (DUBs) has largely relied on covalent complexation of the DUB through its catalytic cysteine with a Ub C-terminal electrophile. The Ub electrophiles are accessed through intein chemistry in conjunction with chemical synthesis. Here, it was asked whether DUB – Ub covalent complexes could instead be accessed by simpler disulfide chemistry using a Ub cysteine mutant in which the last glycine has been replaced with a cysteine. The Ub cysteine mutant displayed a wide variability in disulfide formation across a panel of eukaryotic and prokaryot...
Source: Acta Crystallographica Section D - October 25, 2023 Category: Biochemistry Authors: Negron Teron, K.I. Das, C. Tags: deubiquitinases – ubiquitin complexes disulfide-linked protein complexes research papers Source Type: research
Structural and functional characterization of the novel endo- α (1,4)-fucoidanase Mef1 from the marine bacterium Muricauda eckloniae
Fucoidanases (EC 3.2.1. – ) catalyze the hydrolysis of glycosidic bonds between fucose residues in fucoidans. Fucoidans are a compositionally and structurally diverse class of fucose-containing sulfated polysaccharides that are primarily found in brown seaweeds. Here, the structural characterization of a novel endo- α (1,4)-fucoidanase, Mef1, from the marine bacterium Muricauda eckloniae is presented, showing sequence similarity to members of glycoside hydrolase family 107. Using carbohydrate polyacrylamide gel electrophoresis and nuclear magnetic resonance analyses, it is shown that the fucoidanase Mef1 catalyzes the ...
Source: Acta Crystallographica Section D - October 25, 2023 Category: Biochemistry Authors: Mikkelsen, M.D. Tran, V.H.N. Meier, S. Nguyen, T.T. Holck, J. Cao, H.T.T. Van, T.T.T. Thinh, P.D. Meyer, A.S. Morth, J.P. Tags: fucoidanases GH107 crystal structure ( β / α )8-barrel Ca2 site Mef1 Muricauda eckloniae research papers Source Type: research
Anaerobic fixed-target serial crystallography using sandwiched silicon nitride membranes
This study presents a methodology employing silicon nitride chips arranged in a `sandwich' configuration, enabling reliable room-temperature data collection from oxygen-sensitive samples. The method involves the utilization of a custom-made 3D-printed assembling tool and a MX sample holder. To validate the effectiveness of the proposed method, deoxyhemoglobin and methemoglobin samples were investigated using the BioMAX X-ray macromolecular crystallography beamline, the Balder X-ray absorption spectroscopy beamline and UV – Vis absorption spectroscopy. (Source: Acta Crystallographica Section D)
Source: Acta Crystallographica Section D - October 20, 2023 Category: Biochemistry Authors: Bjel č i ć , M. Sigfridsson Clauss, K.G.V. Aurelius, O. Milas, M. Nan, J. Ursby, T. Tags: fixed-target serial crystallography sample supports sandwiched silicon nitride membranes anaerobic data collection hemoglobin research papers Source Type: research
Structural analysis of wild-type and Val120Thr mutant Candida boidinii formate dehydrogenase by X-ray crystallography
Candida boidinii NAD+-dependent formate dehydrogenase (CbFDH) has gained significant attention for its potential application in the production of biofuels and various industrial chemicals from inorganic carbon dioxide. The present study reports the atomic X-ray crystal structures of wild-type CbFDH at cryogenic and ambient temperatures, as well as that of the Val120Thr mutant at cryogenic temperature, determined at the Turkish Light Source `Turkish DeLight'. The structures reveal new hydrogen bonds between Thr120 and water molecules in the active site of the mutant CbFDH, suggesting increased stability of the active site a...
Source: Acta Crystallographica Section D - October 20, 2023 Category: Biochemistry Authors: Gul, M. Yuksel, B. Bulut, H. DeMirci, H. Tags: formate dehydrogenases Candida boidinii protein engineering X-ray crystallography structural biology structural dynamics Turkish Light Source Turkish DeLight research papers Source Type: research
The impact of molecular variants, crystallization conditions and the space group on ligand – protein complexes: a case study on bacterial phosphotriesterase
A bacterial phosphotriesterase was employed as an experimental paradigm to examine the effects of multiple factors, such as the molecular constructs, the ligands used during protein expression and purification, the crystallization conditions and the space group, on the visualization of molecular complexes of ligands with a target enzyme. In this case, the ligands used were organophosphates that are fragments of the nerve agents and insecticides on which the enzyme acts as a bioscavenger. 12 crystal structures of various phosphotriesterase constructs obtained by directed evolution were analyzed, with resolutions of up to 1....
Source: Acta Crystallographica Section D - October 20, 2023 Category: Biochemistry Authors: Dym, O. Aggarwal, N. Ashani, Y. Leader, H. Albeck, S. Unger, T. Hamer-Rogotner, S. Silman, I. Tawfik, D.S. Sussman, J.L. Tags: crystallization conditions zinc ions phosphotriesterases organophosphates space groups bioscavengers expression systems research papers Source Type: research
Crystal structures of the DExH-box RNA helicase DHX9
DHX9 is a DExH-box RNA helicase with versatile functions in transcription, translation, RNA processing and regulation of DNA replication. DHX9 has recently emerged as a promising target for oncology, but to date no mammalian structures have been published. Here, crystal structures of human, dog and cat DHX9 bound to ADP are reported. The three mammalian DHX9 structures share identical structural folds. Additionally, the overall architecture and the individual domain structures of DHX9 are highly conserved with those of MLE, the Drosophila orthologue of DHX9 previously solved in complex with RNA and a transition-state analo...
Source: Acta Crystallographica Section D - October 20, 2023 Category: Biochemistry Authors: Lee, Y.-T. Sickmier, E.A. Grigoriu, S. Castro, J. Boriack-Sjodin, P.A. Tags: DHX9 RNA helicases DExH box ADP research papers Source Type: research
Domain structure and cross-linking in a giant adhesin from the Mobiluncus mulieris bacterium
Cell-surface proteins known as adhesins enable bacteria to colonize particular environments, and in Gram-positive bacteria often contain autocatalytically formed covalent intramolecular cross-links. While investigating the prevalence of such cross-links, a remarkable example was discovered in Mobiluncus mulieris, a pathogen associated with bacterial vaginosis. This organism encodes a putative adhesin of 7651 residues. Crystallography and mass spectrometry of two selected domains, and AlphaFold structure prediction of the remainder of the protein, were used to show that this adhesin belongs to the family of thioester, isope...
Source: Acta Crystallographica Section D - October 20, 2023 Category: Biochemistry Authors: Young, P.G. Paynter, J.M. Wardega, J.K. Middleditch, M.J. Payne, L.S. Baker, E.N. Squire, C.J. Tags: bacterial adhesins Ig-like domains intramolecular cross-links cell adhesion research papers Source Type: research
Atypical homodimerization revealed by the structure of the (S)-enantioselective haloalkane dehalogenase DmmarA from Mycobacterium marinum
Haloalkane dehalogenases (HLDs) are a family of α / β -hydrolase fold enzymes that employ SN2 nucleophilic substitution to cleave the carbon – halogen bond in diverse chemical structures, the biological role of which is still poorly understood. Atomic-level knowledge of both the inner organization and supramolecular complexation of HLDs is thus crucial to understand their catalytic and noncatalytic functions. Here, crystallographic structures of the (S)-enantioselective haloalkane dehalogenase DmmarA from the waterborne pathogenic microbe Mycobacterium marinum were determined at 1.6 and 1.85 Å resolution. The stru...
Source: Acta Crystallographica Section D - October 20, 2023 Category: Biochemistry Authors: Snajdarova, K. Marques, S.M. Damborsky, J. Bednar, D. Marek, M. Tags: haloalkane dehalogenases Mycobacterium marinum DmmarA homodimerization surface loops enantioselectivity X-ray crystallography SAXS research papers Source Type: research
A user-friendly plug-and-play cyclic olefin copolymer-based microfluidic chip for room-temperature, fixed-target serial crystallography
Over the past two decades, serial X-ray crystallography has enabled the structure determination of a wide range of proteins. With the advent of X-ray free-electron lasers (XFELs), ever-smaller crystals have yielded high-resolution diffraction and structure determination. A crucial need to continue advancement is the efficient delivery of fragile and micrometre-sized crystals to the X-ray beam intersection. This paper presents an improved design of an all-polymer microfluidic `chip' for room-temperature fixed-target serial crystallography that can be tailored to broadly meet the needs of users at either synchrotron or XFEL ...
Source: Acta Crystallographica Section D - September 25, 2023 Category: Biochemistry Authors: Liu, Z. Gu, K.K. Shelby, M.L. Gilbile, D. Lyubimov, A.Y. Russi, S. Cohen, A.E. Narayanasamy, S.R. Botha, S. Kupitz, C. Sierra, R.G. Poitevin, F. Gilardi, A. Lisova, S. Coleman, M.A. Frank, M. Kuhl, T.L. Tags: X-ray crystallography synchrotrons XFELs sample delivery fixed targets cyclic olefin copolymer microfluidics serial crystallography room-temperature SFX research papers Source Type: research
Increasing the bulk of the 1TEL – target linker and retaining the 10 × His tag in a 1TEL – CMG2-vWa construct improves crystal order and diffraction limits
TELSAM-fusion crystallization has the potential to become a revolutionary tool for the facile crystallization of proteins. TELSAM fusion can increase the crystallization rate and enable crystallization at low protein concentrations, in some cases with minimal crystal contacts [Nawarathnage et al. (2022), Open Biol. 12, 210271]. Here, requirements for the linker composition between 1TEL and a fused CMG2 vWa domain were investigated. Ala-Ala, Ala-Val, Thr-Val and Thr-Thr linkers were evaluated, comparing metrics for crystallization propensity and crystal order. The effect on crystallization of removing or retaining the purif...
Source: Acta Crystallographica Section D - September 25, 2023 Category: Biochemistry Authors: Gajjar, P.L. Pedroza Romo, M.J. Litchfield, C.M. Callahan, M. Redd, N. Nawarathnage, S. Soleimani, S. Averett, J. Wilson, E. Lewis, A. Stewart, C. Tseng, Y.-J. Doukov, T. Lebedev, A. Moody, J.D. Tags: polymer-mediated protein crystallization best practices 1TEL TELSAM CMG2 ANTXR2 vWa domains polymer flipping research papers Source Type: research
