Acta crystallographica. Section F, Structural biology communications最新文献

Fucoidan is a complex, sulfated polysaccharide primarily found in brown algae, where it plays important structural and protective roles. Due to its abundance in marine ecosystems, many marine bacteria have evolved diverse and specialized enzymatic systems to degrade fucoidan, although the functions and structures of many of these enzymes remain uncharacterized. Here, we describe the structure of a newly identified fucosidase, FucWf4, which cleaves terminal, unsulfated fucose residues from linear, sulfated fucoidan. FucWf4 does not belong to any known glycoside hydrolase (GH) family, but shows the greatest similarity to GH29 fucosidases. We present the first crystal structure of FucWf4 in complex with fucose, revealing a unique C-terminal domain that resembles a carbohydrate-binding module, although it may have lost its carbohydrate-binding capacity and is absent from canonical GH29 enzymes. Docking experiments suggest the presence of a −1 subsite containing a potential sulfate-binding pocket, which may underlie the substrate specificity of the enzyme. Furthermore, sequence analysis of FucWf4 homologs reveals two distinct clades, likely corresponding to functionally divergent groups. Together, these findings provide new insights into the molecular basis of fucoidan recognition and degradation by this novel enzyme subfamily, laying the groundwork for future functional and structural studies.

Ketohexokinase (KHK) catalyses the initial step in fructose metabolism, converting the furanose form of d-fructose to fructose 1-phosphate in an ATP-dependent reaction. Given its central role in metabolic pathways, KHK has emerged as a target for pharmacological intervention in the treatment of non-alcoholic fatty liver disease, metabolic syndrome, type 2 diabetes and obesity. KHK exists as two isoforms, A and C, which arise from alternative splicing of exon 3, resulting in a differing 45-amino-acid sequence within the 298-amino-acid primary structure of the enzyme. KHK is a biological homodimer, with each subunit adopting an α/β-fold architecture that interlocks with a β-clasp domain. In the case of KHK-C at least two distinct conformations of the β-clasp domain have been identified, whereas this conformational flexibility had not been observed in KHK-A. Here, X-ray crystallographic structural investigations of unliganded murine KHK-A refined to 1.37 Å resolution revealed the adoption of two conformations similar to those adopted by the human ortholog, suggesting that this structural feature is conserved across species. The functional significance of these conformational changes in KHK-A is of particular interest as this isoform has been implicated in cancer metastasis through a `moonlighting' protein kinase activity. Understanding the mechanistic role of conformational shifts in KHK-A may provide insights into its broader physiological functions and therapeutic potential.

Ease of access to data, tools and models expedites scientific research. In structural biology there are now numerous open repositories of experimental and simulated data sets. Being able to easily access and utilize these is crucial to allow researchers to make optimal use of their research effort. The tools presented here are useful for collating existing public cryoEM data sets and/or creating new synthetic cryoEM data sets to aid the development of novel data processing and interpretation algorithms. In recent years, structural biology has seen the development of a multitude of machine-learning-based algorithms to aid numerous steps in the processing and reconstruction of experimental data sets and the use of these approaches has become widespread. Developing such techniques in structural biology requires access to large data sets, which can be cumbersome to curate and unwieldy to make use of. In this paper, we present a suite of Python software packages, which we collectively refer to as PERC (profet, EMPIARreader and CAKED). These are designed to reduce the burden which data curation places upon structural biology research. The protein structure fetcher (profet) package allows users to conveniently download and cleave sequences or structures from the Protein Data Bank or AlphaFold databases. EMPIARreader allows lazy loading of Electron Microscopy Public Image Archive data sets in a machine-learning-compatible structure. The Class Aggregator for Key Electron-microscopy Data (CAKED) package is designed to seamlessly facilitate the training of machine-learning models on electron microscopy data, including electron-cryo-microscopy-specific data augmentation and labeling. These packages may be utilized independently or as building blocks in workflows. All are available in open-source repositories and designed to be easily extensible to facilitate more advanced workflows if required.

The focused issue on the SAMPREP workshop is introduced. The virtual issue is available at https://journals.iucr.org/special_issues/2025/samprep23/.

The enzyme d-aspartate oxidase (DDO) oxidizes acidic d-amino acids using the coenzyme flavin adenine dinucleotide to generate the corresponding α-keto acids and ammonia. DDO differs from d-amino-acid oxidase (DAAO), which acts on neutral and basic d-amino acids. Although the enzymatic properties of DDO have been characterized in several species, the structure of DDO had remained unclear. The structure of DDO derived from Cryptococcus humicola strain UJ1 (chDDO) was determined by X-ray crystallography at 1.70 Å resolution. While the three-dimensional structures of DAAOs are known to be homodimers, chDDO forms a homotetramer. This difference was found to be caused by the deletion of one loop and the insertion of two loops.

Bacillus subtilis DegQ is a 46-amino-acid regulatory protein involved in the DegS–DegU two-component system. DegQ promotes the phosphorylation of DegU by DegS, switching the function of DegU from competence to the induction of poly-γ-glutamate production. To elucidate its structural role, we determined the crystal structures of wild-type DegQ and its mutant DegQS25L. Each DegQ monomer folds into a single α-helix, and four monomers assemble into a tetramer characterized by a four-helix coiled-coil structure. Within the tetramer, two adjacent helices are oriented in the same direction, while the other two are oriented oppositely, forming a pseudo-twofold symmetric arrangement. The mutant form displays disrupted symmetry due to altered helix packing, which is caused by shifts in the coiled-coil heptad register induced by the mutation. Structural predictions using AlphaFold3 suggest that DegQ likely binds to the N-terminal helix bundle of DegS, either as a dimer or as individual monomers. These findings provide structural insight into DegQ oligomerization and its potential role in modulating DegS autophosphorylation and DegU binding.

Several cyanobacterial species, including Crocosphaera subtropica ATCC 51142, accumulate cyanobacterial starch instead of glycogen, although nearly all cyanobacteria accumulate glycogen. The glycogen-producing Synechococcus elongatus PCC 7942 possesses one α-glucan phosphorylase (Pho) isozyme, whereas strain 51142 has three Pho isozymes. Based on their primary structures, these enzymes belong to glycosyl transferase (GT) family 35, with the cyanobacterial GT35-type Phos further subdivided into types I–III. In this study, to elucidate the significance of the coexistence of multiple GT35-type Pho isozymes, those from strain 51142 (type I, cce_1629; type II, cce_1603 and cce_5186) and strain 7942 (type I, Synpcc7942_0244) were overexpressed in Escherichia coli and biochemically characterized. All isozymes catalysed the phosphorolysis and reverse phosphorolysis reactions. The type I isozyme from a cyanobacterial starch-producing strain (cce_1629) differed in substrate specificity and specific activity compared with the others. The behaviour towards the effectors (AMP and ATP) of the type I and type II isozymes differed from each other. These findings enhance our understanding of the roles of cyanobacterial Pho isozymes in α-glucan metabolism. Furthermore, recombinant cce_1603 was crystallized using the hanging-drop vapour-diffusion method. Crystals were obtained at 293 K in the presence of 10 mM maltoheptaose, 45%(w/v) PEG 400, 0.1 M Tris–HCl pH 8.0, 0.2 M lithium sulfate. The crystals belonged to space group R32 (hexagonal setting) with unit-cell parameters a = b = 267.23, c = 204.43 Å, and diffracted to beyond 2.70 Å resolution. Matthews coefficient calculations indicated the presence of two molecules in the asymmetric unit. Structural determination is currently under way. The crystal structure of cce_1603 will aid in the understanding of the structural basis of cyanobacterial GT35-type Pho isozymes.

Gasdermin D (GSDMD) is a protein that has gained significant attention in recent years due to its crucial role in inflammatory cell death, particularly pyroptosis. Pyroptosis is a highly inflammatory form of programmed cell death that is triggered by various microbial infections and sterile inflammatory stimuli. GSDMD acts as an executioner molecule in this process, leading to the release of pro-inflammatory cytokines and amplifying the immune response. Here, we present a higher resolution, significantly improved apo crystal structure of the deposited mouse structure model that will be beneficial for structure-based drug-design approaches towards this important pharmacological target.