Journal of Structural Biology: X最新文献

Conformational variability in the D2 loop of Plasmodium Apical Membrane antigen 1 疟原虫顶膜抗原 1 D2 环的构象变异性

IF 3.5

Journal of Structural Biology: X Pub Date : 2024-09-10 DOI: 10.1016/j.yjsbx.2024.100110

{"title":"Conformational variability in the D2 loop of Plasmodium Apical Membrane antigen 1","authors":"","doi":"10.1016/j.yjsbx.2024.100110","DOIUrl":"10.1016/j.yjsbx.2024.100110","url":null,"abstract":"<div><p>Apical Membrane Antigen 1 (AMA1) plays a vital role in the invasion of the host erythrocyte by the malaria parasite, <em>Plasmodium</em>. It is thus an important target for vaccine and anti-malaria therapeutic strategies that block the invasion process. AMA1, present on the surface of the parasite, interacts with RON2, a component of the parasite’s rhoptry neck (RON) protein complex, which is transferred to the erythrocyte membrane during invasion. The D2 loop of AMA1 plays an essential role in invasion as it partially covers the RON2-binding site and must therefore be displaced for invasion to proceed. Several structural studies have shown that the D2 loop is very mobile, a property that is probably important for the function of AMA1. Here we present three crystal structures of AMA1 from <em>P. falciparum</em> (strains 3D7 and FVO) and <em>P. vivax</em> (strain Sal1), in which the D2 loop could be largely traced in the electron density maps. The D2 loop of PfAMA1-FVO and PvAMA1 (as a complex with a monoclonal antibody Fab) has a conformation previously noted in the <em>P. knowlesi</em> AMA1 structure. The D2 loop of PfAMA1-3D7, however, reveals a novel conformation. We analyse the conformational variability of the D2 loop in these structures, together with those previously reported. Three different conformations can be distinguished, all of which are highly helical and show some similarity in their secondary structure organisation. We discuss the significance of these observations in the light of the flexible nature of the D2 loop and its role in AMA1 function.</p></div>","PeriodicalId":17238,"journal":{"name":"Journal of Structural Biology: X","volume":null,"pages":null},"PeriodicalIF":3.5,"publicationDate":"2024-09-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.sciencedirect.com/science/article/pii/S2590152424000151/pdfft?md5=18146cbe19a02e66f922067a1ea42cae&pid=1-s2.0-S2590152424000151-main.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142243547","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}

引用次数: 0

Structure of SARS-CoV-2 MTase nsp14 with the inhibitor STM957 reveals inhibition mechanism that is shared with a poxviral MTase VP39 SARS-CoV-2 MT 酶 nsp14 与抑制剂 STM957 的结构揭示了与痘病毒 MT 酶 VP39 共享的抑制机制

IF 3.5

Journal of Structural Biology: X Pub Date : 2024-07-29 DOI: 10.1016/j.yjsbx.2024.100109

{"title":"Structure of SARS-CoV-2 MTase nsp14 with the inhibitor STM957 reveals inhibition mechanism that is shared with a poxviral MTase VP39","authors":"","doi":"10.1016/j.yjsbx.2024.100109","DOIUrl":"10.1016/j.yjsbx.2024.100109","url":null,"abstract":"<div><p>Nsp14 is an RNA methyltransferase (MTase) encoded by all coronaviruses. In fact, many viral families, including DNA viruses, encode MTases that catalyze the methylation of the RNA precap structure, resulting in fully capped viral RNA. This capping is crucial for efficient viral RNA translation, stability, and immune evasion. Our previous research identified nsp14 inhibitors based on the chemical scaffold of its methyl donor − the S-adenosyl methionine (SAM) − featuring a modified adenine base and a substituted arylsulfonamide. However, the binding mode of these inhibitors was based only on docking experiments. To uncover atomic details of nsp14 inhibition we solved the crystal structure of nsp14 bound to STM957. The structure revealed the atomic details of nsp14 inhibition such that the 7-deaza-adenine moiety of STM957 forms specific interactions with Tyr368, Ala353, and Phe367, while the arylsulfonamide moiety engages with Asn388 and Phe506. The large aromatic substituent at the 7-deaza position displaces a network of water molecules near the adenine base. Surprisingly, this was recently observed in the case of an unrelated monkeypox MTase VP39, where the 7-deaza modified SAH analogs also displaced water molecules from the vicinity of the active site.</p></div>","PeriodicalId":17238,"journal":{"name":"Journal of Structural Biology: X","volume":null,"pages":null},"PeriodicalIF":3.5,"publicationDate":"2024-07-29","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.sciencedirect.com/science/article/pii/S259015242400014X/pdfft?md5=adc7af4fba68360ac9d28aa0d253354d&pid=1-s2.0-S259015242400014X-main.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141962088","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}

引用次数: 0

Do selectivity filter carbonyls in K+ channels flip away from the pore? Two-dimensional infrared spectroscopy study K+ 通道中的选择性过滤羰基是否会从孔中翻转？二维红外光谱研究

IF 3.5

Journal of Structural Biology: X Pub Date : 2024-07-15 DOI: 10.1016/j.yjsbx.2024.100108

{"title":"Do selectivity filter carbonyls in K+ channels flip away from the pore? Two-dimensional infrared spectroscopy study","authors":"","doi":"10.1016/j.yjsbx.2024.100108","DOIUrl":"10.1016/j.yjsbx.2024.100108","url":null,"abstract":"<div><p>Molecular dynamics simulations revealed that the carbonyls of the Val residue in the conserved selectivity filter sequence TVGTG of potassium ion channels can flip away from the pore to form hydrogen bonds with the network of water molecules residing behind the selectivity filter. Such a configuration has been proposed to be relevant for C-type inactivation. Experimentally, X-ray crystallography of the KcsA channel admits the possibility that the Val carbonyls can flip, but it cannot decisively confirm the existence of such a configuration. In this study, we combined molecular dynamics simulations and line shape theory to design two-dimensional infrared spectroscopy experiments that can corroborate the existence of the selectivity filter configuration with flipped Val carbonyls. This ability to distinguish between flipped and unflipped carbonyls is based on the varying strength of the electric field inside and outside the pore, which is directly linked to carbonyl stretching frequencies that can be resolved using infrared spectroscopy.</p></div>","PeriodicalId":17238,"journal":{"name":"Journal of Structural Biology: X","volume":null,"pages":null},"PeriodicalIF":3.5,"publicationDate":"2024-07-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.sciencedirect.com/science/article/pii/S2590152424000138/pdfft?md5=13bf2c257aaf33fbd68799a7ce5f4412&pid=1-s2.0-S2590152424000138-main.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141689647","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}

引用次数: 0

MicroED structure of the C11 cysteine protease clostripain C11 半胱氨酸蛋白酶 Clostripain 的显微电子数据结构

IF 3.5

Journal of Structural Biology: X Pub Date : 2024-07-06 DOI: 10.1016/j.yjsbx.2024.100107

Yasmeen N. Ruma , Guanhong Bu , Johan Hattne , Tamir Gonen

{"title":"MicroED structure of the C11 cysteine protease clostripain","authors":"Yasmeen N. Ruma , Guanhong Bu , Johan Hattne , Tamir Gonen","doi":"10.1016/j.yjsbx.2024.100107","DOIUrl":"https://doi.org/10.1016/j.yjsbx.2024.100107","url":null,"abstract":"<div><p>Clostripain secreted from <em>Clostridium histolyticum</em> is the founding member of the C11 family of Clan CD cysteine peptidases, which is an important group of peptidases secreted by numerous bacteria. Clostripain is an arginine-specific endopeptidase. Because of its efficacy as a cysteine peptidase, it is widely used in laboratory settings. Despite its importance the structure of clostripain remains unsolved. Here we describe the first structure of an active form of <em>C. histolyticum</em> clostripain determined at 2.5 Å resolution using microcrystal electron diffraction (MicroED). The structure was determined from a single nanocrystal after focused ion beam milling. The structure of clostripain shows a typical Clan CD α/β/α sandwich architecture and the Cys231/His176 catalytic dyad in the active site. It has a large electronegative substrate binding pocket showing its ability to accommodate large and diverse substrates. A loop in the heavy chain formed between residues 452 and 457 is potentially important for substrate binding. In conclusion, this result demonstrates the importance of MicroED to determine the unknown structure of macromolecules such as clostripain, which can be further used as a platform to study substrate binding and design of potential inhibitors against this class of peptidases.</p></div>","PeriodicalId":17238,"journal":{"name":"Journal of Structural Biology: X","volume":null,"pages":null},"PeriodicalIF":3.5,"publicationDate":"2024-07-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.sciencedirect.com/science/article/pii/S2590152424000126/pdfft?md5=42cb6b31866b2698d485367c031389f5&pid=1-s2.0-S2590152424000126-main.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141594445","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}

引用次数: 0

Nucleic acid-binding KH domain proteins influence a spectrum of biological pathways including as part of membrane-localized complexes 核酸结合 KH 结构域蛋白影响一系列生物通路，包括作为膜定位复合物的一部分发挥作用

IF 3.5

Journal of Structural Biology: X Pub Date : 2024-06-27 DOI: 10.1016/j.yjsbx.2024.100106

Md Kamrul Hasan , L. Jeannine Brady

{"title":"Nucleic acid-binding KH domain proteins influence a spectrum of biological pathways including as part of membrane-localized complexes","authors":"Md Kamrul Hasan , L. Jeannine Brady","doi":"10.1016/j.yjsbx.2024.100106","DOIUrl":"https://doi.org/10.1016/j.yjsbx.2024.100106","url":null,"abstract":"<div><p>K-Homology domain (KH domain) proteins bind single-stranded nucleic acids, influence protein–protein interactions of proteins that harbor them, and are found in all kingdoms of life. In concert with other functional protein domains KH domains contribute to a variety of critical biological activities, often within higher order machineries including membrane-localized protein complexes. Eukaryotic KH domain proteins are linked to developmental processes, morphogenesis, and growth regulation, and their aberrant expression is often associated with cancer. Prokaryotic KH domain proteins are involved in integral cellular activities including cell division and protein translocation. Eukaryotic and prokaryotic KH domains share structural features, but are differentiated based on their structural organizations. In this review, we explore the structure/function relationships of known examples of KH domain proteins, and highlight cases in which they function within or at membrane surfaces. We also summarize examples of KH domain proteins that influence bacterial virulence and pathogenesis. We conclude the article by discussing prospective research avenues that could be pursued to better investigate this largely understudied protein category.</p></div>","PeriodicalId":17238,"journal":{"name":"Journal of Structural Biology: X","volume":null,"pages":null},"PeriodicalIF":3.5,"publicationDate":"2024-06-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.sciencedirect.com/science/article/pii/S2590152424000114/pdfft?md5=0e422cb348e848d0c60e6c2df8352de4&pid=1-s2.0-S2590152424000114-main.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141486539","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}

引用次数: 0

Accurate size-based protein localization from cryo-ET tomograms 从低温电子断层扫描图中准确定位蛋白质大小

IF 3.5

Journal of Structural Biology: X Pub Date : 2024-06-26 DOI: 10.1016/j.yjsbx.2024.100104

Weisheng Jin , Ye Zhou , Alberto Bartesaghi

{"title":"Accurate size-based protein localization from cryo-ET tomograms","authors":"Weisheng Jin , Ye Zhou , Alberto Bartesaghi","doi":"10.1016/j.yjsbx.2024.100104","DOIUrl":"https://doi.org/10.1016/j.yjsbx.2024.100104","url":null,"abstract":"<div><p>Cryo-electron tomography (cryo-ET) combined with sub-tomogram averaging (STA) allows the determination of protein structures imaged within the native context of the cell at near-atomic resolution. Particle picking is an essential step in the cryo-ET/STA image analysis pipeline that consists in locating the position of proteins within crowded cellular tomograms so that they can be aligned and averaged in 3D to improve resolution. While extensive work in 2D particle picking has been done in the context of single-particle cryo-EM, comparatively fewer strategies have been proposed to pick particles from 3D tomograms, in part due to the challenges associated with working with noisy 3D volumes affected by the missing wedge. While strategies based on 3D template-matching and deep learning are commonly used, these methods are computationally expensive and require either an external template or manual labelling which can bias the results and limit their applicability. Here, we propose a size-based method to pick particles from tomograms that is fast, accurate, and does not require external templates or user provided labels. We compare the performance of our approach against a commonly used algorithm based on deep learning, crYOLO, and show that our method: i) has higher detection accuracy, ii) does not require user input for labeling or time-consuming training, and iii) runs efficiently on non-specialized CPU hardware. We demonstrate the effectiveness of our approach by automatically detecting particles from tomograms representing different types of samples and using these particles to determine the high-resolution structures of ribosomes imaged <em>in vitro</em> and <em>in situ</em>.</p></div>","PeriodicalId":17238,"journal":{"name":"Journal of Structural Biology: X","volume":null,"pages":null},"PeriodicalIF":3.5,"publicationDate":"2024-06-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.sciencedirect.com/science/article/pii/S2590152424000096/pdfft?md5=9bf56ad8420d48d82b350489a5b2005c&pid=1-s2.0-S2590152424000096-main.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141594447","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}

引用次数: 0

Solution structure, dynamics and tetrahedral assembly of Anti-TRAP, a homo-trimeric triskelion-shaped regulator of tryptophan biosynthesis in Bacillus subtilis 枯草芽孢杆菌中色氨酸生物合成的同源三叉戟形调控因子 Anti-TRAP 的溶液结构、动力学和四面体组装

IF 3.5

Journal of Structural Biology: X Pub Date : 2024-06-11 DOI: 10.1016/j.yjsbx.2024.100103

Craig A. McElroy , Elihu C. Ihms , Deepak Kumar Yadav , Melody L. Holmquist , Vibhuti Wadhwa , Vicki H. Wysocki , Paul Gollnick , Mark P. Foster

{"title":"Solution structure, dynamics and tetrahedral assembly of Anti-TRAP, a homo-trimeric triskelion-shaped regulator of tryptophan biosynthesis in Bacillus subtilis","authors":"Craig A. McElroy , Elihu C. Ihms , Deepak Kumar Yadav , Melody L. Holmquist , Vibhuti Wadhwa , Vicki H. Wysocki , Paul Gollnick , Mark P. Foster","doi":"10.1016/j.yjsbx.2024.100103","DOIUrl":"https://doi.org/10.1016/j.yjsbx.2024.100103","url":null,"abstract":"<div><p>Cellular production of tryptophan is metabolically expensive and tightly regulated. The small <em>Bacillus subtilis</em> zinc binding Anti-TRAP protein (AT), which is the product of the <em>yczA/rtpA</em> gene, is upregulated in response to accumulating levels of uncharged tRNA<sup>Trp</sup> through a T-box antitermination mechanism. AT binds to the undecameric axially symmetric ring-shaped protein TRAP (<em>trp</em> RNA Binding Attenuation Protein), thereby preventing it from binding to the <em>trp</em> leader RNA. This reverses the inhibitory effect of TRAP on transcription and translation of the <em>trp</em> operon. AT principally adopts two symmetric oligomeric states, a trimer (AT<sub>3</sub>) featuring three-fold axial symmetry or a dodecamer (AT<sub>12</sub>) comprising a tetrahedral assembly of trimers, whereas only the trimeric form binds and inhibits TRAP. We apply native mass spectrometry (nMS) and small-angle x-ray scattering (SAXS), together with analytical ultracentrifugation (AUC) to monitor the pH and concentration-dependent equilibrium between the trimeric and dodecameric structural forms of AT. In addition, we use solution nuclear magnetic resonance (NMR) spectroscopy to determine the solution structure of AT<sub>3</sub>, while heteronuclear <sup>15</sup>N relaxation measurements on both oligomeric forms of AT provide insights into the dynamic properties of binding-active AT<sub>3</sub> and binding-inactive AT<sub>12</sub>, with implications for TRAP binding and inhibition.</p></div>","PeriodicalId":17238,"journal":{"name":"Journal of Structural Biology: X","volume":null,"pages":null},"PeriodicalIF":3.5,"publicationDate":"2024-06-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.sciencedirect.com/science/article/pii/S2590152424000084/pdfft?md5=571b0da5dc58532b76dc49e84cbcb4d5&pid=1-s2.0-S2590152424000084-main.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141486538","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}

引用次数: 0

Eliminating the missing cone challenge through innovative approaches 通过创新方法消除缺锥难题

IF 2.9

Journal of Structural Biology: X Pub Date : 2024-06-01 DOI: 10.1016/j.yjsbx.2024.100102

Cody Gillman , Guanhong Bu , Emma Danelius , Johan Hattne , Brent L. Nannenga , Tamir Gonen

{"title":"Eliminating the missing cone challenge through innovative approaches","authors":"Cody Gillman , Guanhong Bu , Emma Danelius , Johan Hattne , Brent L. Nannenga , Tamir Gonen","doi":"10.1016/j.yjsbx.2024.100102","DOIUrl":"https://doi.org/10.1016/j.yjsbx.2024.100102","url":null,"abstract":"<div><p>Microcrystal electron diffraction (MicroED) has emerged as a powerful technique for unraveling molecular structures from microcrystals too small for X-ray diffraction. However, a significant hurdle arises with plate-like crystals that consistently orient themselves flat on the electron microscopy grid. If the normal of the plate correlates with the axes of the crystal lattice, the crystal orientations accessible for measurement are restricted because the crystal cannot be arbitrarily rotated. This limits the information that can be acquired, resulting in a missing cone of information. We recently introduced a novel crystallization strategy called suspended drop crystallization and proposed that crystals in a suspended drop could effectively address the challenge of preferred crystal orientation. Here we demonstrate the success of the suspended drop approach in eliminating the missing cone in two samples that crystallize as thin plates: bovine liver catalase and the SARS‑CoV‑2 main protease (Mpro). This innovative solution proves indispensable for crystals exhibiting systematic preferred orientations, unlocking new possibilities for structure determination by MicroED.</p></div>","PeriodicalId":17238,"journal":{"name":"Journal of Structural Biology: X","volume":null,"pages":null},"PeriodicalIF":2.9,"publicationDate":"2024-06-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.sciencedirect.com/science/article/pii/S2590152424000072/pdfft?md5=7540527cb3411fb26f5be3a6bf21bfcd&pid=1-s2.0-S2590152424000072-main.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141290118","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}

引用次数: 0

Coordination of bilayer properties by an inward-rectifier K+ channel is a cooperative process driven by protein-lipid interaction 内向整流 K+ 通道对双分子层特性的协调是一个由蛋白质-脂质相互作用驱动的合作过程

IF 2.9

Journal of Structural Biology: X Pub Date : 2024-06-01 DOI: 10.1016/j.yjsbx.2024.100101

Evan J. van Aalst , Maryam Yekefallah , Roy A. M. van Beekveld , Eefjan Breukink , Markus Weingarth , Benjamin J. Wylie

{"title":"Coordination of bilayer properties by an inward-rectifier K+ channel is a cooperative process driven by protein-lipid interaction","authors":"Evan J. van Aalst , Maryam Yekefallah , Roy A. M. van Beekveld , Eefjan Breukink , Markus Weingarth , Benjamin J. Wylie","doi":"10.1016/j.yjsbx.2024.100101","DOIUrl":"https://doi.org/10.1016/j.yjsbx.2024.100101","url":null,"abstract":"<div><p>Physical properties of biological membranes directly or indirectly govern biological processes. Yet, the interplay between membrane and integral membrane proteins is difficult to assess due to reciprocal effects between membrane proteins, individual lipids, and membrane architecture. Using solid-state NMR (SSNMR) we previously showed that KirBac1.1, a bacterial Inward-Rectifier K<sup>+</sup> channel, nucleates bilayer ordering and microdomain formation through tethering anionic lipids. Conversely, these lipids cooperatively bind cationic residues to activate the channel and initiate K<sup>+</sup> flux. The mechanistic details governing the relationship between cooperative lipid loading and bilayer ordering are, however, unknown. To investigate, we generated KirBac1.1 samples with different concentrations of <sup>13</sup>C-lableded phosphatidyl glycerol (PG) lipids and acquired a full suite of SSNMR 1D temperature series experiments using the ordered all-trans (AT) and disordered <em>trans</em>-gauche (TG) acyl conformations as markers of bilayer dynamics. We observed increased AT ordered signal, decreased TG disordered signal, and increased bilayer melting temperature with increased PG concentration. Further, we identified cooperativity between ordering and direct binding of PG lipids, indicating KirBac1.1-driven bilayer ordering and microdomain formation is a classically cooperative Hill-type process driven by and predicated upon direct binding of PG lipids. Our results provide unique mechanistic insight into how proteins and lipids in tandem contribute to supramolecular bilayer heterogeneity in the lipid membrane.</p></div>","PeriodicalId":17238,"journal":{"name":"Journal of Structural Biology: X","volume":null,"pages":null},"PeriodicalIF":2.9,"publicationDate":"2024-06-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.sciencedirect.com/science/article/pii/S2590152424000060/pdfft?md5=93c4cf2e0eae744a1e20dcd4448f8094&pid=1-s2.0-S2590152424000060-main.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141240761","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}

引用次数: 0

Optimizing NMR fragment-based drug screening for membrane protein targets 优化基于核磁共振片段的膜蛋白靶点药物筛选

IF 2.9

Journal of Structural Biology: X Pub Date : 2024-06-01 DOI: 10.1016/j.yjsbx.2024.100100

Geoffrey C. Li , Manuel A. Castro , Thilini Ukwaththage, Charles R. Sanders

{"title":"Optimizing NMR fragment-based drug screening for membrane protein targets","authors":"Geoffrey C. Li , Manuel A. Castro , Thilini Ukwaththage, Charles R. Sanders","doi":"10.1016/j.yjsbx.2024.100100","DOIUrl":"https://doi.org/10.1016/j.yjsbx.2024.100100","url":null,"abstract":"<div><p>NMR spectroscopy has played a pivotal role in fragment-based drug discovery by coupling detection of weak ligand-target binding with structural mapping of the binding site. Fragment-based screening by NMR has been successfully applied to many soluble protein targets, but only to a limited number of membrane proteins, despite the fact that many drug targets are membrane proteins. This is partly because of difficulties preparing membrane proteins for NMR—especially human membrane proteins—and because of the inherent complexity associated with solution NMR spectroscopy on membrane protein samples, which require the inclusion of membrane-mimetic agents such as micelles, nanodiscs, or bicelles. Here, we developed a generalizable protocol for fragment-based screening of membrane proteins using NMR. We employed two human membrane protein targets, both in fully protonated detergent micelles: the single-pass C-terminal domain of the amyloid precursor protein, C99, and the tetraspan peripheral myelin protein 22 (PMP22). For both we determined the optimal NMR acquisition parameters, protein concentration, protein-to-micelle ratio, and upper limit to the concentration of D<sub>6</sub>-DMSO in screening samples. Furthermore, we conducted preliminary screens of a plate-format molecular fragment mixture library using our optimized conditions and were able to identify hit compounds that selectively bound to the respective target proteins. It is hoped that the approaches presented here will be useful in complementing existing methods for discovering lead compounds that target membrane proteins.</p></div>","PeriodicalId":17238,"journal":{"name":"Journal of Structural Biology: X","volume":null,"pages":null},"PeriodicalIF":2.9,"publicationDate":"2024-06-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.sciencedirect.com/science/article/pii/S2590152424000059/pdfft?md5=39c55f7648d09ecd7660ace75e9b5e22&pid=1-s2.0-S2590152424000059-main.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141240760","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}

引用次数: 0