Structures of l-proline trans-hydroxylase reveal the catalytic specificity and provide deeper insight into AKG-dependent hydroxylation
l-Proline hydroxylase is a member of the non-heme Fe2+/ α -ketoglutarate (AKG)-dependent hydroxylase family that catalyzes the reaction from l-proline to hydroxy-l-proline, which is widely used in drug synthesis, biochemistry, food supplementation and cosmetic industries. Here, the first crystal structure of l-proline trans-hydroxylase and its complexes with substrate and product are reported, which reveal the structural basis of trans – cis proline hydroxylation selectivity. Structure comparison with other AKG-dependent hydroxylases identifies conserved amino acid residues, which may serve as signatures of in-line or o...
Source: Acta Crystallographica Section D - March 28, 2023 Category: Biochemistry Authors: Hu, X. Huang, X. Liu, J. Zheng, P. Gong, W. Yang, L. Tags: AKG-dependent hydroxylases hydroxy-l-proline l-proline trans-hydroxylases – cis proline hydroxylation selectivity catalytic specificity AKG binding modes research papers Source Type: research
Ambient temperature structure of phosphoketolase from Bifidobacterium longum determined by serial femtosecond X-ray crystallography
Phosphoketolase and transketolase are thiamine diphosphate-dependent enzymes and play a central role in the primary metabolism of bifidobacteria: the bifid shunt. The enzymes both catalyze phosphorolytic cleavage of xylulose 5-phosphate or fructose 6-phosphate in the first reaction step, but possess different substrate specificity in the second reaction step, where phosphoketolase and transketolase utilize inorganic phosphate (Pi) and d-ribose 5-phosphate, respectively, as the acceptor substrate. Structures of Bifidobacterium longum phosphoketolase holoenzyme and its complex with a putative inhibitor, phosphoenolpyruvate, ...
Source: Acta Crystallographica Section D - March 28, 2023 Category: Biochemistry Authors: Nakata, K. Kashiwagi, T. Kunishima, N. Naitow, H. Matsuura, Y. Miyano, H. Mizukoshi, T. Tono, K. Yabashi, M. Nango, E. Iwata, S. Tags: enzyme mechanisms enzyme inhibitors phosphoketolases thiamine diphosphate-dependent enzymes bifid shunt serial femtosecond crystallography X-ray free-electron laser research papers Source Type: research
Likelihood-based docking of models into cryo-EM maps
Optimized docking of models into cryo-EM maps requires exploiting an understanding of the signal expected in the data to minimize the calculation time while maintaining sufficient signal. The likelihood-based rotation function used in crystallography can be employed to establish plausible orientations in a docking search. A phased likelihood translation function yields scores for the placement and rigid-body refinement of oriented models. Optimized strategies for choices of the resolution of data from the cryo-EM maps to use in the calculations and the size of search volumes are based on expected log-likelihood-gain scores...
Source: Acta Crystallographica Section D - March 15, 2023 Category: Biochemistry Authors: Mill á n, C. McCoy, A.J. Terwilliger, T.C. Read, R.J. Tags: likelihood cryo-EM docking information gain research papers Source Type: research
Likelihood-based signal and noise analysis for docking of models into cryo-EM maps
Fast, reliable docking of models into cryo-EM maps requires understanding of the errors in the maps and the models. Likelihood-based approaches to errors have proven to be powerful and adaptable in experimental structural biology, finding applications in both crystallography and cryo-EM. Indeed, previous crystallographic work on the errors in structural models is directly applicable to likelihood targets in cryo-EM. Likelihood targets in Fourier space are derived here to characterize, based on the comparison of half-maps, the direction- and resolution-dependent variation in the strength of both signal and noise in the data...
Source: Acta Crystallographica Section D - March 15, 2023 Category: Biochemistry Authors: Read, R.J. Mill á n, C. McCoy, A.J. Terwilliger, T.C. Tags: likelihood cryo-EM docking information gain research papers Source Type: research
A tetramerization domain in prokaryotic and eukaryotic transcription regulators homologous to p53
Transcriptional regulation usually requires the action of several proteins that either repress or activate a promotor of an open reading frame. These proteins can counteract each other, thus allowing tight regulation of the transcription of the corresponding genes, where tight repression is often linked to DNA looping or cross-linking. Here, the tetramerization domain of the bacterial gene repressor Rco from Bacillus subtilis plasmid pLS20 (RcopLS20) has been identified and its structure is shown to share high similarity to the tetramerization domain of the well known p53 family of human tumor suppressors, despite lacking ...
Source: Acta Crystallographica Section D - March 1, 2023 Category: Biochemistry Authors: Bernardo, N. Crespo, I. Cuppari, A. Meijer, W.J.J. Boer, D.R. Tags: transcription regulation DNA looping tumor suppression structural biology research papers Source Type: research
Structural basis of DNA binding by YdaT, a functional equivalent of the CII repressor in the cryptic prophage CP-933P from Escherichia coli O157:H7
YdaT is a functional equivalent of the CII repressor in certain lambdoid phages and prophages. YdaT from the cryptic prophage CP-933P in the genome of Escherichia coli O157:H7 is functional as a DNA-binding protein and recognizes a 5 ′ -TTGATTN6AATCAA-3 ′ inverted repeat. The DNA-binding domain is a helix – turn – helix (HTH)-containing POU domain and is followed by a long α -helix ( α 6) that forms an antiparallel four-helix bundle, creating a tetramer. The loop between helix α 2 and the recognition helix α 3 in the HTH motif is unusually long compared with typical HTH motifs, and is highly variable in sequenc...
Source: Acta Crystallographica Section D - February 27, 2023 Category: Biochemistry Authors: Proli č -Kalin š ek, M. Volkov, A.N. Had ž i, S. Van Dyck, J. Bervoets, I. Charlier, D. Loris, R. Tags: CII repressors DNA binding YdaT transcription regulation toxin – antitoxin research papers Source Type: research
Accelerating crystal structure determination with iterative AlphaFold prediction
Experimental structure determination can be accelerated with artificial intelligence (AI)-based structure-prediction methods such as AlphaFold. Here, an automatic procedure requiring only sequence information and crystallographic data is presented that uses AlphaFold predictions to produce an electron-density map and a structural model. Iterating through cycles of structure prediction is a key element of this procedure: a predicted model rebuilt in one cycle is used as a template for prediction in the next cycle. This procedure was applied to X-ray data for 215 structures released by the Protein Data Bank in a recent six-m...
Source: Acta Crystallographica Section D - February 27, 2023 Category: Biochemistry Authors: Terwilliger, T.C. Afonine, P.V. Liebschner, D. Croll, T.I. McCoy, A.J. Oeffner, R.D. Williams, C.J. Poon, B.K. Richardson, J.S. Read, R.J. Adams, P.D. Tags: AlphaFold artificial intelligence automated structure determination model building research papers Source Type: research
Correction of rhodopsin serial crystallography diffraction intensities for a lattice-translocation defect
Rhodopsin is a G-protein-coupled receptor that detects light and initiates the intracellular signalling cascades that underpin vertebrate vision. Light sensitivity is achieved by covalent linkage to 11-cis retinal, which isomerizes upon photo-absorption. Serial femtosecond crystallography data collected from rhodopsin microcrystals grown in the lipidic cubic phase were used to solve the room-temperature structure of the receptor. Although the diffraction data showed high completeness and good consistency to 1.8 Å resolution, prominent electron-density features remained unaccounted for throughout the unit cell after mo...
Source: Acta Crystallographica Section D - February 27, 2023 Category: Biochemistry Authors: Rodrigues, M.J. Casadei, C.M. Weinert, T. Panneels, V. Schertler, G.F.X. Tags: serial crystallography lattice-translocation defects membrane proteins lipidic cubic phase G protein-coupled receptors research papers Source Type: research
Obtaining anomalous and ensemble information from protein crystals from 220 K up to physiological temperatures
X-ray crystallography has been invaluable in delivering structural information about proteins. Previously, an approach has been developed that allows high-quality X-ray diffraction data to be obtained from protein crystals at and above room temperature. Here, this previous work is built on and extended by showing that high-quality anomalous signal can be obtained from single protein crystals using diffraction data collected at 220 K up to physiological temperatures. The anomalous signal can be used to directly determine the structure of a protein, i.e. to phase the data, as is routinely performed under cryoconditions. ...
Source: Acta Crystallographica Section D - February 27, 2023 Category: Biochemistry Authors: Doukov, T. Herschlag, D. Yabukarski, F. Tags: native phasing anomalous diffraction room temperature data collection research papers Source Type: research
Errors in structural biology are not the exception
During the COVID-19 pandemic, the structural biology community swung into action quickly and efficiently, and many urgent questions were solved by macromolecular structure determination. The Coronavirus Structural Task Force evaluated all structures from SARS-CoV-1 and SARS-CoV-2, but errors in measurement, data processing and modelling are present beyond these structures and throughout the structures deposited in the Protein Data Bank. Identifying them is only the first step; in order to minimize the impact that errors have in structural biology, error culture needs to change. It should be emphasized that the atomic model...
Source: Acta Crystallographica Section D - February 27, 2023 Category: Biochemistry Authors: Gao, Y. Thorn, V. Thorn, A. Tags: errors structural biology modelling quality indicators validation scientific comment Source Type: research
20 years of crystal hits: progress and promise in ultrahigh-throughput crystallization screening
Diffraction-based structural methods contribute a large fraction of the biomolecular structural models available, providing a critical understanding of macromolecular architecture. These methods require crystallization of the target molecule, which remains a primary bottleneck in crystal-based structure determination. The National High-Throughput Crystallization Center at Hauptman – Woodward Medical Research Institute has focused on overcoming obstacles to crystallization through a combination of robotics-enabled high-throughput screening and advanced imaging to increase the success of finding crystallization conditions....
Source: Acta Crystallographica Section D - February 27, 2023 Category: Biochemistry Authors: Lynch, M.L. Snell, M.E. Potter, S.A. Snell, E.H. Bowman, S.E.J. Tags: macromolecular crystallization macromolecular crystallography crystallization facilities high-throughput crystallization crystallization screening feature articles Source Type: research
2023 update of template tables for reporting biomolecular structural modelling of small-angle scattering data
In 2017, guidelines were published for reporting structural modelling of small-angle scattering (SAS) data from biomolecules in solution that exemplified best-practice documentation of experiments and analysis. Since then, there has been significant progress in SAS data and model archiving, and the IUCr journal editors announced that the IUCr biology journals will require the deposition of SAS data used in biomolecular structure solution into a public archive, as well as adherence to the 2017 reporting guidelines. In this context, the reporting template tables accompanying the 2017 publication guidelines have been revie...
Source: Acta Crystallographica Section D - February 6, 2023 Category: Biochemistry Authors: Trewhella, J. Jeffries, C.M. Whitten, A.E. Tags: small-angle scattering SAXS SANS contrast variation biomolecular structural modelling template tables research papers Source Type: research
Structural and functional characterization of a thermostable secretory phospholipase A2 from Sciscionella marina and its application in liposome biotransformation
Secretory phospholipase A2 (sPLA2), which hydrolyzes the sn-2 acyl bond of lecithin in a Ca2+-dependent manner, is an important enzyme in the oil and oleochemical industries. However, most sPLA2s are not stable under process conditions. Therefore, a thermostable sPLA2 was investigated in this study. A marine bacterial sPLA2 isolated from Sciscionella marina (Sm-sPLA2) was catalytically active even after 5 h of incubation at high temperatures of up to 50 ° C, which is outstanding compared with a representative bacterial sPLA2 (i.e. sPLA2 from Streptomyces violaceoruber; Sv-sPLA2). Consistent with this, the melting temp...
Source: Acta Crystallographica Section D - February 6, 2023 Category: Biochemistry Authors: Kang, B.-G. Kwon, S.-Y. Lee, H.-R. Hwang, Y. Youn, S.-Y. Oh, C. Park, J.-B. Cha, S.-S. Tags: phospholipase A2 Sciscionella marina thermostability crystal structure loop anchoring liposome biotransformation research papers Source Type: research
The LH – DH module of bacterial replicative helicases is the common binding site for DciA and other helicase loaders
During the initiation step of bacterial genome replication, replicative helicases depend on specialized proteins for their loading onto oriC. DnaC and DnaI were the first loaders to be characterized. However, most bacteria do not contain any of these genes, which are domesticated phage elements that have replaced the ancestral and unrelated loader gene dciA several times during evolution. To understand how DciA assists the loading of DnaB, the crystal structure of the complex from Vibrio cholerae was determined, in which two VcDciA molecules interact with a dimer of VcDnaB without changing its canonical structure. The data...
Source: Acta Crystallographica Section D - February 6, 2023 Category: Biochemistry Authors: Cargemel, C. Marsin, S. Noiray, M. Legrand, P. Bounoua, H. Li de la Sierra-Gallay, I. Walbott, H. Quevillon-Cheruel, S. Tags: DnaB · DciA complex crystal structure helicase loaders bacterial replicative helicases interaction modules replicative helicase hijacking Vibrio cholerae research papers Source Type: research
Crystal structure of a polyglycine hydrolase determined using a RoseTTAFold model
The objective of this research was to investigate the proteolytic mechanism and other characteristics by structural and biochemical means. Here, the first atomic structure of a polyglycine hydrolase was identified. It was solved by X-ray crystallography using a RoseTTAFold model, taking advantage of recent technical advances in structure prediction. PGHs are composed of two domains: the N- and C-domains. The N-domain is a novel tertiary fold with an as-yet unknown function that is found across all kingdoms of life. The C-domain shares structural similarities with class C β -lactamases, including a common catalytic nucleop...
Source: Acta Crystallographica Section D - February 6, 2023 Category: Biochemistry Authors: Dowling, N.V. Naumann, T.A. Price, N.P.J. Rose, D.R. Tags: RoseTTAFold fungi chitinase-modifying proteins polyglycine hydrolases β -lactamases research papers Source Type: research
