Spirochetes are a widely existing group of bacteria with a distinct morphology. Some spirochetes are important human pathogens that utilize periplasmic flagella to achieve motility and host infection. The motors that drive the rotation of periplasmic flagella have a unique spirochete-specific feature, termed the collar, crucial for the flat-wave morphology and motility of the Lyme disease spirochete Borrelia burgdorferi.
View Article and Find Full Text PDFWhile novel deep learning and statistics-based techniques predict accurate structural models for proteins and non-coding RNA, describing their macromolecular conformations in solution is still challenging. Small-angle X-ray scattering (SAXS) in solution is an efficient technique to validate structural predictions by comparing the experimental SAXS profile with those calculated from predicted structures. There are two main challenges in comparing SAXS profiles to RNA structures: the structures often lack cations necessary for stability and charge neutralization, and a single structure inadequately represents the conformational plasticity of RNA.
View Article and Find Full Text PDFAdvanced deep learning and statistical methods can predict structural models for RNA molecules. However, RNAs are flexible, and it remains difficult to describe their macromolecular conformations in solutions where varying conditions can induce conformational changes. Small-angle x-ray scattering (SAXS) in solution is an efficient technique to validate structural predictions by comparing the experimental SAXS profile with those calculated from predicted structures.
View Article and Find Full Text PDFStaphylococcus aureus expresses three high-affinity neutrophil serine protease (NSP) inhibitors known as the extracellular adherence protein domain (EAPs) proteins. Whereas EapH1 and EapH2 are comprised of a single EAP domain, the modular extracellular adherence protein (Eap) from S. aureus strain Mu50 consists of four EAP domains.
View Article and Find Full Text PDFGlycosylation is a predominant strategy plants use to fine-tune the properties of small molecule metabolites to affect their bioactivity, transport, and storage. It is also important in biotechnology and medicine as many glycosides are utilized in human health. Small molecule glycosylation is largely carried out by family 1 glycosyltransferases.
View Article and Find Full Text PDFActivating signal co-integrator complex 1 (ASCC1) acts with ASCC-ALKBH3 complex in alkylation damage responses. ASCC1 uniquely combines two evolutionarily ancient domains: nucleotide-binding K-Homology (KH) (associated with regulating splicing, transcriptional, and translation) and two-histidine phosphodiesterase (PDE; associated with hydrolysis of cyclic nucleotide phosphate bonds). Germline mutations link loss of ASCC1 function to spinal muscular atrophy with congenital bone fractures 2 (SMABF2).
View Article and Find Full Text PDFThe transition of IgA antibodies into clinical development is crucial because they have the potential to create a new class of therapeutics with superior pathogen neutralization, cancer cell killing, and immunomodulation capacity compared to IgG. However, the biological role of IgA glycans in these processes needs to be better understood. This study provides a detailed biochemical, biophysical, and structural characterization of recombinant monomeric human IgA2, which varies in the amount/locations of attached glycans.
View Article and Find Full Text PDFElectron bifurcation (BF) is an evolutionarily ancient energy coupling mechanism in anaerobes, whose associated enzymatic machinery remains enigmatic. In BF-flavoenzymes, a chemically high-potential electron forms in a thermodynamically favorable fashion by simultaneously dropping the potential of a second electron before its donation to physiological acceptors. The cryo-EM and spectroscopic analyses of the BF-enzyme Fix/EtfABCX from suggest that the BF-site contains a special flavin-adenine dinucleotide and, upon its reduction with NADH, a low-potential electron transfers to ferredoxin and a high-potential electron reduces menaquinone.
View Article and Find Full Text PDFThe function of cellobiose dehydrogenase (CDH) in biosensors, biofuel cells, and as a physiological redox partner of lytic polysaccharide monooxygenase (LPMO) is based on its role as an electron donor. Before donating electrons to LPMO or electrodes, an interdomain electron transfer from the catalytic FAD-containing dehydrogenase domain to the electron shuttling cytochrome domain of CDH is required. This study investigates the role of two crucial amino acids located at the dehydrogenase domain on domain interaction and interdomain electron transfer by structure-based engineering.
View Article and Find Full Text PDFThe recent clinical and commercial success of lipid nanoparticles (LNPs) for nucleic acid delivery has incentivized the development of new technologies to manufacture LNPs. As new technologies emerge, researchers must determine which technologies to assess and how to perform comparative evaluations. In this article, we use a quality-by-design approach to systematically investigate how the mixer technology used to form LNPs influences LNPstructure.
View Article and Find Full Text PDFThe interdomain electron transfer (IET) between the catalytic flavodehydrogenase domain and the electron-transferring cytochrome domain of cellobiose dehydrogenase (CDH) plays an essential role in biocatalysis, biosensors and biofuel cells, as well as in its natural function as an auxiliary enzyme of lytic polysaccharide monooxygenase. We investigated the mobility of the cytochrome and dehydrogenase domains of CDH, which is hypothesised to limit IET in solution by small angle X-ray scattering (SAXS). CDH from Myriococcum thermophilum (syn.
View Article and Find Full Text PDFWith three FDA-approved products, lipid nanoparticles (LNPs) are under intensive development for delivering wide-ranging nucleic acid therapeutics. A significant challenge for LNP development is insufficient understanding of structure-activity relationship (SAR). Small changes in chemical composition and process parameters can affect LNP structure, significantly impacting performance and .
View Article and Find Full Text PDFLipid nanoparticles (LNPs) have been widely investigated for nucleic acid therapeutic delivery, and demonstrated their potential in enabling new mRNA vaccines. LNPs are usually formulated with multi-lipid components and the composition variables may impact their structural properties. Here, we investigated the impact of helper lipids on physicochemical properties of LNPs using a Design of Experiments (DoE) definitive screening design.
View Article and Find Full Text PDFRhamnogalacturonan I (RGI) is a structurally complex pectic polysaccharide with a backbone of alternating rhamnose and galacturonic acid residues substituted with arabinan and galactan side chains. Galactan synthase 1 (GalS1) transfers galactose and arabinose to either extend or cap the β-1,4-galactan side chains of RGI, respectively. Here we report the structure of GalS1 from Populus trichocarpa, showing a modular protein consisting of an N-terminal domain that represents the founding member of a new family of carbohydrate-binding module, CBM95, and a C-terminal glycosyltransferase family 92 (GT92) catalytic domain that adopts a GT-A fold.
View Article and Find Full Text PDFZNF410 is a highly-conserved transcription factor, remarkable in that it recognizes a 15-base pair DNA element but has just a single responsive target gene in mammalian erythroid cells. ZNF410 includes a tandem array of five zinc-fingers (ZFs), surrounded by uncharacterized N- and C-terminal regions. Unexpectedly, full-length ZNF410 has reduced DNA binding affinity, compared to that of the isolated DNA binding ZF array, both in vitro and in cells.
View Article and Find Full Text PDFAccurate protein structure predictions, enabled by recent advances in machine learning algorithms, provide an entry point to probing structural mechanisms and to integrating and querying many types of biochemical and biophysical results. Limitations in such protein structure predictions can be reduced and addressed through comparison to experimental Small Angle X-ray Scattering (SAXS) data that provides protein structural information in solution. SAXS data can not only validate computational predictions, but can improve conformational and assembly prediction to produce atomic models that are consistent with solution data and biologically relevant states.
View Article and Find Full Text PDFSmall-angle X-ray Scattering (SAXS) has been a versatile technique for studying biomolecules in solution for several decades now. Developments in SAXS techniques that integrate in situ purification with a high-throughput, multimodal design philosophy have revolutionized the reach and tempo of BioSAXS experiments. The current zenith of the field comes in the form of size exclusion chromatography coupled SAXS with in-line multiangle light scattering (SEC-SAXS-MALS).
View Article and Find Full Text PDFThrough an expansive international effort that involved data collection on 12 small-angle X-ray scattering (SAXS) and four small-angle neutron scattering (SANS) instruments, 171 SAXS and 76 SANS measurements for five proteins (ribonuclease A, lysozyme, xylanase, urate oxidase and xylose isomerase) were acquired. From these data, the solvent-subtracted protein scattering profiles were shown to be reproducible, with the caveat that an additive constant adjustment was required to account for small errors in solvent subtraction. Further, the major features of the obtained consensus SAXS data over the q measurement range 0-1 Å are consistent with theoretical prediction.
View Article and Find Full Text PDFLipid nanoparticles (LNPs) are gaining traction in the field of nucleic acid delivery following the success of two mRNA vaccines against COVID-19. As one of the constituent lipids on LNP surfaces, PEGylated lipids (PEG-lipids) play an important role in defining LNP physicochemical properties and biological interactions. Previous studies indicate that LNP performance is modulated by tuning PEG-lipid parameters including PEG size and architecture, carbon tail type and length, as well as the PEG-lipid molar ratio in LNPs.
View Article and Find Full Text PDFRegulation of bacteriophage gene expression involves repressor proteins that bind and downregulate early lytic promoters. A large group of mycobacteriophages code for repressors that are unusual in also terminating transcription elongation at numerous binding sites (stoperators) distributed across the phage genome. Here we provide the X-ray crystal structure of a mycobacteriophage immunity repressor bound to DNA, which reveals the binding of a monomer to an asymmetric DNA sequence using two independent DNA binding domains.
View Article and Find Full Text PDFThe RAS-RAF pathway is one of the most commonly dysregulated in human cancers. Despite decades of study, understanding of the molecular mechanisms underlying dimerization and activation of the kinase RAF remains limited. Recent structures of inactive RAF monomer and active RAF dimer bound to 14-3-3 have revealed the mechanisms by which 14-3-3 stabilizes both RAF conformations via specific phosphoserine residues.
View Article and Find Full Text PDFBackground: COBRA™ (COnditional Bispecific Redirected Activation) T-cell engagers are designed to target solid tumors as a single polypeptide chain prodrug that becomes activated by proteolysis in the tumor microenvironment. One COBRA molecule comprises seven Ig domains: three single-domain antibodies (sdAbs) recognizing a tumor target or human serum albumin (HSA), and CD3ε-binding variable fragment heavy chain (VH) and variable fragment light chain (VL) and their inactivated counterparts, VHi and VLi. Pairing of VH and VL, and VLi and VHi into single-chain variable fragments (Fv) is prevented by shortened inter-domain linkers.
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