Journal of Structural Biology: X最新文献

{"title":"Structural plasticity in the loop region of engineered lipocalins with novel ligand specificities, so-called Anticalins","authors":"S. Achatz,&nbsp;A. Jarasch,&nbsp;A. Skerra","doi":"10.1016/j.yjsbx.2021.100054","DOIUrl":"10.1016/j.yjsbx.2021.100054","url":null,"abstract":"<div><p>Anticalins are generated via combinatorial protein design on the basis of the lipocalin protein scaffold and constitute a novel class of small and robust engineered binding proteins that offer prospects for applications in medical therapy as well as <em>in vivo</em> diagnostics as an alternative to antibodies. The lipocalins are natural binding proteins with diverse ligand specificities which share a simple architecture with a central eight-stranded antiparallel β-barrel and an α-helix attached to its side. At the open end of the β-barrel, four structurally variable loops connect the β-strands in a pair-wise manner and, together, shape the ligand pocket. Using targeted random mutagenesis in combination with molecular selection techniques, this loop region can be reshaped to generate pockets for the tight binding of various ligands ranging from small molecules over peptides to proteins. While such Anticalin proteins can be derived from different natural lipocalins, the human lipocalin 2 (Lcn2) scaffold proved particularly successful for the design of binding proteins with novel specificities and, over the years, more than 20 crystal structures of Lcn2-based Anticalins have been elucidated. In this graphical structural biology review we illustrate the conformational variability that emerged in the loop region of these functionally diverse artificial binding proteins in comparison with the natural scaffold. Our present analysis provides picturesque evidence of the high structural plasticity around the binding site of the lipocalins which explains the proven tolerance toward excessive mutagenesis, thus demonstrating remarkable resemblance to the complementarity-determining region of antibodies (immunoglobulins).</p></div>","PeriodicalId":17238,"journal":{"name":"Journal of Structural Biology: X","volume":"6 ","pages":"Article 100054"},"PeriodicalIF":2.9,"publicationDate":"2022-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/d9/65/main.PMC8693463.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"39789119","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Subtomogram analysis: The sum of a tomogram’s particles reveals molecular structure in situ","authors":"Friedrich Förster","doi":"10.1016/j.yjsbx.2022.100063","DOIUrl":"https://doi.org/10.1016/j.yjsbx.2022.100063","url":null,"abstract":"<div><p>Cryo-electron tomography is uniquely suited to provide insights into the molecular architecture of cells and tissue in the native state. While frozen hydrated specimens tolerate sufficient electron doses to distinguish different types of particles in a tomogram, the accumulating beam damage does not allow resolving their detailed molecular structure individually. Statistical methods for subtomogram averaging and classification that coherently enhance the signal of particles corresponding to copies of the same type of macromolecular allow obtaining much higher resolution insights into macromolecules. Here, I review the developments in subtomogram analysis at Wolfgang Baumeister’s laboratory that make the dream of structural biology in the native cell become reality.</p></div>","PeriodicalId":17238,"journal":{"name":"Journal of Structural Biology: X","volume":"6 ","pages":"Article 100063"},"PeriodicalIF":2.9,"publicationDate":"2022-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.sciencedirect.com/science/article/pii/S2590152422000046/pdfft?md5=609a521d1a590ad494462d2a057cfdd3&pid=1-s2.0-S2590152422000046-main.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"72075253","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Subtomogram averaging for biophysical analysis and supramolecular context","authors":"Lauren Ann Metskas ,&nbsp;Rosalie Wilfong ,&nbsp;Grant J. Jensen","doi":"10.1016/j.yjsbx.2022.100076","DOIUrl":"10.1016/j.yjsbx.2022.100076","url":null,"abstract":"<div><p>Recent advances in hardware, software and computing power have led to increasingly ambitious applications of cryo-electron tomography and subtomogram averaging. It is now possible to reveal both structures and biophysical relationships like protein binding partners and small molecule occupancy in these experiments. However, some data processing choices require the user to prioritize structure or biophysical context. Here, we present a modified subtomogram averaging approach that preserves both capabilities. By increasing the accuracy of particle-picking, performing alignment and averaging on all subtomograms, and decreasing reliance on symmetry and tight masks, the usability of tomography and subtomogram averaging data for biophysical analyses is greatly increased without negatively impacting structural refinements.</p></div>","PeriodicalId":17238,"journal":{"name":"Journal of Structural Biology: X","volume":"6 ","pages":"Article 100076"},"PeriodicalIF":2.9,"publicationDate":"2022-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/01/ea/main.PMC9596874.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10644661","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Biological solid-state NMR: Integrative across different scientific disciplines","authors":"Marc Baldus","doi":"10.1016/j.yjsbx.2022.100075","DOIUrl":"10.1016/j.yjsbx.2022.100075","url":null,"abstract":"<div><p>For almost five decades, solid-state NMR (ssNMR) has been used to study complex biomolecular systems. This article gives a view on how ssNMR methods and applications have evolved during this time period in a broader structural biology context. It also discusses possible directions for additional developments and the future role of ssNMR in a life science context and beyond.</p></div>","PeriodicalId":17238,"journal":{"name":"Journal of Structural Biology: X","volume":"6 ","pages":"Article 100075"},"PeriodicalIF":2.9,"publicationDate":"2022-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9523391/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"40389445","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Does dentine mineral change with anatomical location, microscopic site and patient age?","authors":"Arosha T. Weerakoon ,&nbsp;Crystal Cooper ,&nbsp;Ian A. Meyers ,&nbsp;Nicholas Condon ,&nbsp;Christopher Sexton ,&nbsp;David Thomson ,&nbsp;Pauline J. Ford ,&nbsp;Anne L. Symons","doi":"10.1016/j.yjsbx.2022.100060","DOIUrl":"10.1016/j.yjsbx.2022.100060","url":null,"abstract":"<div><h3>Objective</h3><p>To determine the effect of patient age (young or mature), anatomical location (shallow/deep and central/peripheral) and microscopic site (intertubular/peritubular) on dentine mineral density, distribution and composition.</p></div><div><h3>Methods</h3><p>Extracted posterior teeth from young (aged 19–20 years, N = 4) and mature (aged 54–77 years, N = 4) subjects were prepared to shallow and deep slices. The dentine surface elemental composition was investigated in a SEM using Backscattered Electron (BSE) micrographs, Energy Dispersive X-ray Spectroscopy, and Integrated Mineral Analysis. Qualitative comparisons and quantitative measures using machine learning were used to analyse the BSE images. Quantitative outcomes were compared using quantile or linear regression models with bootstrapping to account for the multiple measures per sample. Subsequently, a Xenon Plasma Focussed Ion Beam Scanning Electron Microscopy (Xe PFIB-SEM) was used to mill large area (100 µm) cross-sections to investigate morphology through the dentine tubules using high resolution secondary electron micrographs.</p></div><div><h3>Results</h3><p>With age, dentine mineral composition remains stable, but density changes with anatomical location and microscopic site. Microscopically, accessory tubules spread into intertubular dentine (ITD) from the main tubule lumens. Within the lumens, mineral deposits form calcospherites in the young that eventually coalesce in mature tubules and branches. The mineral occlusion in mature dentine increases overall ITD density to reflect peritubular dentine (PTD) infiltrate. The ITD observed in micrographs remained consistent for age and observation plane to suggest tubule deposition affects overall dentine density. Mineral density depends on the relative distribution of PTD to ITD that varies with anatomical location.</p></div><div><h3>Significance</h3><p>Adhesive materials may interact differently within a tooth as well as in different age groups.</p></div>","PeriodicalId":17238,"journal":{"name":"Journal of Structural Biology: X","volume":"6 ","pages":"Article 100060"},"PeriodicalIF":2.9,"publicationDate":"2022-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/91/16/main.PMC8818708.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"39613919","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Multimodal imaging reveals membrane skeleton reorganisation during reticulocyte maturation and differences in dimple and rim regions of mature erythrocytes","authors":"Adam J. Blanch ,&nbsp;Juan Nunez-Iglesias ,&nbsp;Arman Namvar ,&nbsp;Sebastien Menant ,&nbsp;Oliver Looker ,&nbsp;Vijay Rajagopal ,&nbsp;Wai-Hong Tham ,&nbsp;Leann Tilley ,&nbsp;Matthew W.A. Dixon","doi":"10.1016/j.yjsbx.2021.100056","DOIUrl":"10.1016/j.yjsbx.2021.100056","url":null,"abstract":"<div><p>The red blood cell (RBC) is remarkable in its ability to deform as it passages through the vasculature. Its deformability derives from a spectrin-actin protein network that supports the cell membrane and provides strength and flexibility, however questions remain regarding the assembly and maintenance of the skeletal network. Using scanning electron microscopy (SEM) and atomic force microscopy (AFM) we have examined the nanoscale architecture of the cytoplasmic side of membrane discs prepared from reticulocytes and mature RBCs. Immunofluorescence microscopy was used to probe the distribution of spectrin and other membrane skeleton proteins. We found that the cell surface area decreases by up to 30% and the spectrin-actin network increases in density by approximately 20% as the reticulocyte matures. By contrast, the inter-junctional distance and junctional density increase only by 3–4% and 5–9%, respectively. This suggests that the maturation-associated reduction in surface area is accompanied by an increase in spectrin self-association to form higher order oligomers. We also examined the mature RBC membrane in the edge (rim) and face (dimple) regions of mature RBCs and found the rim contains about 1.5% more junctional complexes compared to the dimple region. A 2% increase in band 4.1 density in the rim supports these structural measurements.</p></div>","PeriodicalId":17238,"journal":{"name":"Journal of Structural Biology: X","volume":"6 ","pages":"Article 100056"},"PeriodicalIF":2.9,"publicationDate":"2022-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/80/3d/main.PMC8688873.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"39780147","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"In-cell DNP NMR reveals multiple targeting effect of antimicrobial peptide","authors":"Frances Separovic ,&nbsp;Vinzenz Hofferek ,&nbsp;Anthony P. Duff ,&nbsp;Malcom J. McConville ,&nbsp;Marc-Antoine Sani","doi":"10.1016/j.yjsbx.2022.100074","DOIUrl":"10.1016/j.yjsbx.2022.100074","url":null,"abstract":"<div><p>Dynamic nuclear polarization NMR spectroscopy was used to investigate the effect of the antimicrobial peptide (AMP) maculatin 1.1 on <em>E. coli</em> cells. The enhanced ¹⁵N NMR signals from nucleic acids, proteins and lipids identified a number of unanticipated physiological responses to peptide stress, revealing that membrane-active AMPs can have a multi-target impact on <em>E. coli</em> cells. DNP-enhanced ¹⁵N-observed ³¹P-dephased REDOR NMR allowed monitoring how Mac1 induced DNA condensation and prevented intermolecular salt bridges between the main <em>E. coli</em> lipid phosphatidylethanolamine (PE) molecules. The latter was supported by similar results obtained using <em>E. coli</em> PE lipid systems. Overall, the ability to monitor the action of antimicrobial peptides <em>in situ</em> will provide greater insight into their mode of action.</p></div>","PeriodicalId":17238,"journal":{"name":"Journal of Structural Biology: X","volume":"6 ","pages":"Article 100074"},"PeriodicalIF":2.9,"publicationDate":"2022-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/2c/60/main.PMC9486116.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"33478817","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Recent advances in the structural biology of encapsulin bacterial nanocompartments","authors":"Timothy Wiryaman ,&nbsp;Navtej Toor","doi":"10.1016/j.yjsbx.2022.100062","DOIUrl":"10.1016/j.yjsbx.2022.100062","url":null,"abstract":"<div><p>Large capsid-like nanocompartments called encapsulins are common in bacteria and archaea and contain cargo proteins with diverse functions. Advances in cryo-electron microscopy have enabled structure determination of many encapsulins in recent years. Here we summarize findings from recent encapsulin structures that have significant implications for their biological roles. We also compare important features such as the E-loop, cargo-peptide binding site, and the fivefold axis channel in different structures. In addition, we describe the discovery of a flavin-binding pocket within the encapsulin shell that may reveal a role for this nanocompartment in iron metabolism.</p></div>","PeriodicalId":17238,"journal":{"name":"Journal of Structural Biology: X","volume":"6 ","pages":"Article 100062"},"PeriodicalIF":2.9,"publicationDate":"2022-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/6d/f3/main.PMC8802124.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10643654","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Colloid assembly and transformation (CAT): The relationship of PILP to biomineralization","authors":"Laurie Gower,&nbsp;Jeremy Elias","doi":"10.1016/j.yjsbx.2021.100059","DOIUrl":"10.1016/j.yjsbx.2021.100059","url":null,"abstract":"<div><p>The field of biomineralization has undergone a revolution in the past 25 years, which paralleled the discovery by Gower of a polymer-induced liquid-precursor (PILP) mineralization process. She proposed this <em>in vitro</em> model system might be useful for studying the role biopolymers play in biomineralization; however, the ramifications of this pivotal discovery were slow to be recognized. This was presumably because it utilized simple polypeptide additives, and at that time it was not recognized that the charged proteins intimately associated with biominerals are often intrinsically disordered proteins (IDPs). Over the years, many enigmatic biomineral features have been emulated with this model system, too many to be mere coincidence. Yet the PILP system continues to be underacknowledged, probably because of its namesake, which indicates a “liquid precursor”, while we now know the phase appears to have viscoelastic character. Another factor is the confusing semantics that arose from the discovery of multiple “non-classical crystallization” pathways. This review suggests a more relevant terminology for the polymer-modulated reactions is “colloid assembly and transformation (CAT)”, which we believe more accurately captures the key stages involved in both biomineralization and the PILP process. The PILP model system has helped to decipher the key role that biopolymers, namely the IDPs, play in modulating biomineralization processes, which was not readily accomplished in living biological systems. Some remaining challenges in understanding the organic–inorganic interactions involved in biomineralization are discussed, which further highlight how the PILP model system may prove invaluable for studying the simple, yet complex, CAT crystallization pathway.</p></div>","PeriodicalId":17238,"journal":{"name":"Journal of Structural Biology: X","volume":"6 ","pages":"Article 100059"},"PeriodicalIF":2.9,"publicationDate":"2022-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/4a/88/main.PMC8749173.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"39688133","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Hydrogens and hydrogen-bond networks in macromolecular MicroED data","authors":"Max T.B. Clabbers ,&nbsp;Michael W. Martynowycz ,&nbsp;Johan Hattne ,&nbsp;Tamir Gonen","doi":"10.1016/j.yjsbx.2022.100078","DOIUrl":"10.1016/j.yjsbx.2022.100078","url":null,"abstract":"<div><p>Microcrystal electron diffraction (MicroED) is a powerful technique utilizing electron cryo-microscopy (cryo-EM) for protein structure determination of crystalline samples too small for X-ray crystallography. Electrons interact with the electrostatic potential of the sample, which means that the scattered electrons carry information about the charged state of atoms and provide relatively stronger contrast for visualizing hydrogen atoms. Accurately identifying the positions of hydrogen atoms, and by extension the hydrogen bonding networks, is of importance for understanding protein structure and function, in particular for drug discovery. However, identification of individual hydrogen atom positions typically requires atomic resolution data, and has thus far remained elusive for macromolecular MicroED. Recently, we presented the <em>ab initio</em> structure of triclinic hen egg-white lysozyme at 0.87 Å resolution. The corresponding data were recorded under low exposure conditions using an electron-counting detector from thin crystalline lamellae. Here, using these subatomic resolution MicroED data, we identified over a third of all hydrogen atom positions based on strong difference peaks, and directly visualize hydrogen bonding interactions and the charged states of residues. Furthermore, we find that the hydrogen bond lengths are more accurately described by the inter-nuclei distances than the centers of mass of the corresponding electron clouds. We anticipate that MicroED, coupled with ongoing advances in data collection and refinement, can open further avenues for structural biology by uncovering the hydrogen atoms and hydrogen bonding interactions underlying protein structure and function.</p></div>","PeriodicalId":17238,"journal":{"name":"Journal of Structural Biology: X","volume":"6 ","pages":"Article 100078"},"PeriodicalIF":2.9,"publicationDate":"2022-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9731847/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10779799","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}