Pub Date : 2024-05-03DOI: 10.1107/S2053230X24003868
Yi Xue, Zhen Wu, Xue Kang
The Rib domain, which is often found as tandem-repeat structural modules in surface proteins of Gram-positive bacteria, plays important roles in mediating interactions of bacteria with their environments and hosts. A comprehensive structural analysis of various Rib domains is essential to fully understand their impact on the structure and functionality of these bacterial adhesins. To date, structural information has been limited for this expansive group of domains. In this study, the high-resolution crystal structure of the second member of the long Rib domain, a unique subclass within the Rib-domain family, derived from Limosilactobacillus reuteri is presented. The data not only demonstrate a highly conserved structure within the long Rib domain, but also highlight an evolutionary convergence in structural architecture with other modular domains found in cell-adhesion molecules.
{"title":"Crystal structure of the long Rib domain of the LPXTG-anchored surface protein from Limosilactobacillus reuteri","authors":"Yi Xue, Zhen Wu, Xue Kang","doi":"10.1107/S2053230X24003868","DOIUrl":"10.1107/S2053230X24003868","url":null,"abstract":"<p>The Rib domain, which is often found as tandem-repeat structural modules in surface proteins of Gram-positive bacteria, plays important roles in mediating interactions of bacteria with their environments and hosts. A comprehensive structural analysis of various Rib domains is essential to fully understand their impact on the structure and functionality of these bacterial adhesins. To date, structural information has been limited for this expansive group of domains. In this study, the high-resolution crystal structure of the second member of the long Rib domain, a unique subclass within the Rib-domain family, derived from <i>Limosilactobacillus reuteri</i> is presented. The data not only demonstrate a highly conserved structure within the long Rib domain, but also highlight an evolutionary convergence in structural architecture with other modular domains found in cell-adhesion molecules.</p>","PeriodicalId":7029,"journal":{"name":"Acta crystallographica. Section F, Structural biology communications","volume":null,"pages":null},"PeriodicalIF":0.9,"publicationDate":"2024-05-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140857368","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-05-01DOI: 10.1107/S2053230X24003911
Guilherme Vilela-Alves, Rita Rebelo Manuel, Neide Pedrosa, Inês A Cardoso Pereira, Maria João Romão, Cristiano Mota
Molybdenum- or tungsten-dependent formate dehydrogenases have emerged as significant catalysts for the chemical reduction of CO2 to formate, with biotechnological applications envisaged in climate-change mitigation. The role of Met405 in the active site of Desulfovibrio vulgaris formate dehydrogenase AB (DvFdhAB) has remained elusive. However, its proximity to the metal site and the conformational change that it undergoes between the resting and active forms suggests a functional role. In this work, the M405S variant was engineered, which allowed the active-site geometry in the absence of methionine Sδ interactions with the metal site to be revealed and the role of Met405 in catalysis to be probed. This variant displayed reduced activity in both formate oxidation and CO2 reduction, together with an increased sensitivity to oxygen inactivation.
{"title":"Structural and biochemical characterization of the M405S variant of Desulfovibrio vulgaris formate dehydrogenase.","authors":"Guilherme Vilela-Alves, Rita Rebelo Manuel, Neide Pedrosa, Inês A Cardoso Pereira, Maria João Romão, Cristiano Mota","doi":"10.1107/S2053230X24003911","DOIUrl":"10.1107/S2053230X24003911","url":null,"abstract":"<p><p>Molybdenum- or tungsten-dependent formate dehydrogenases have emerged as significant catalysts for the chemical reduction of CO<sub>2</sub> to formate, with biotechnological applications envisaged in climate-change mitigation. The role of Met405 in the active site of Desulfovibrio vulgaris formate dehydrogenase AB (DvFdhAB) has remained elusive. However, its proximity to the metal site and the conformational change that it undergoes between the resting and active forms suggests a functional role. In this work, the M405S variant was engineered, which allowed the active-site geometry in the absence of methionine S<sup>δ</sup> interactions with the metal site to be revealed and the role of Met405 in catalysis to be probed. This variant displayed reduced activity in both formate oxidation and CO<sub>2</sub> reduction, together with an increased sensitivity to oxygen inactivation.</p>","PeriodicalId":7029,"journal":{"name":"Acta crystallographica. Section F, Structural biology communications","volume":null,"pages":null},"PeriodicalIF":0.9,"publicationDate":"2024-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11134731/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140849963","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-05-01Epub Date: 2024-05-20DOI: 10.1107/S2053230X2400414X
Matthew P Agdanowski, Roger Castells-Graells, Michael R Sawaya, Duilio Cascio, Todd O Yeates, Mark A Arbing
Imaging scaffolds composed of designed protein cages fused to designed ankyrin repeat proteins (DARPins) have enabled the structure determination of small proteins by cryogenic electron microscopy (cryo-EM). One particularly well characterized scaffold type is a symmetric tetrahedral assembly composed of 24 subunits, 12 A and 12 B, which has three cargo-binding DARPins positioned on each vertex. Here, the X-ray crystal structure of a representative tetrahedral scaffold in the apo state is reported at 3.8 Å resolution. The X-ray crystal structure complements recent cryo-EM findings on a closely related scaffold, while also suggesting potential utility for crystallographic investigations. As observed in this crystal structure, one of the three DARPins, which serve as modular adaptors for binding diverse `cargo' proteins, present on each of the vertices is oriented towards a large solvent channel. The crystal lattice is unusually porous, suggesting that it may be possible to soak crystals of the scaffold with small (≤30 kDa) protein cargo ligands and subsequently determine cage-cargo structures via X-ray crystallography. The results suggest the possibility that cryo-EM scaffolds may be repurposed for structure determination by X-ray crystallography, thus extending the utility of electron-microscopy scaffold designs for alternative structural biology applications.
通过低温电子显微镜(cryo-EM)测定小分子蛋白质的结构,可以利用由设计的蛋白质笼与设计的杏仁蛋白重复蛋白(DARPins)融合而成的成像支架。其中一种特征特别明显的支架类型是由 24 个亚基(12 个 A 和 12 个 B)组成的对称四面体组装体,每个顶点上都有三个与货物结合的 DARPins。在此,我们以 3.8 Å 的分辨率报道了一个具有代表性的四面体支架在 apo 状态下的 X 射线晶体结构。该 X 射线晶体结构补充了最近对一个密切相关支架的低温电子显微镜研究结果,同时也表明了晶体学研究的潜在用途。正如在该晶体结构中所观察到的,存在于每个顶点上的三个 DARPins(作为模块化适配器用于结合各种 "货物 "蛋白)中的一个面向一个大的溶剂通道。晶格异常多孔,这表明有可能用小型(≤30 kDa)蛋白质货物配体浸泡支架晶体,然后通过 X 射线晶体学确定笼-货结构。这些结果表明,低温电子显微镜支架有可能被重新用于通过 X 射线晶体学确定结构,从而将电子显微镜支架设计的用途扩展到其他结构生物学应用领域。
{"title":"X-ray crystal structure of a designed rigidified imaging scaffold in the ligand-free conformation.","authors":"Matthew P Agdanowski, Roger Castells-Graells, Michael R Sawaya, Duilio Cascio, Todd O Yeates, Mark A Arbing","doi":"10.1107/S2053230X2400414X","DOIUrl":"10.1107/S2053230X2400414X","url":null,"abstract":"<p><p>Imaging scaffolds composed of designed protein cages fused to designed ankyrin repeat proteins (DARPins) have enabled the structure determination of small proteins by cryogenic electron microscopy (cryo-EM). One particularly well characterized scaffold type is a symmetric tetrahedral assembly composed of 24 subunits, 12 A and 12 B, which has three cargo-binding DARPins positioned on each vertex. Here, the X-ray crystal structure of a representative tetrahedral scaffold in the apo state is reported at 3.8 Å resolution. The X-ray crystal structure complements recent cryo-EM findings on a closely related scaffold, while also suggesting potential utility for crystallographic investigations. As observed in this crystal structure, one of the three DARPins, which serve as modular adaptors for binding diverse `cargo' proteins, present on each of the vertices is oriented towards a large solvent channel. The crystal lattice is unusually porous, suggesting that it may be possible to soak crystals of the scaffold with small (≤30 kDa) protein cargo ligands and subsequently determine cage-cargo structures via X-ray crystallography. The results suggest the possibility that cryo-EM scaffolds may be repurposed for structure determination by X-ray crystallography, thus extending the utility of electron-microscopy scaffold designs for alternative structural biology applications.</p>","PeriodicalId":7029,"journal":{"name":"Acta crystallographica. Section F, Structural biology communications","volume":null,"pages":null},"PeriodicalIF":0.9,"publicationDate":"2024-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11134730/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141070264","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-04-30DOI: 10.1107/S2053230X2400356X
Xinping Ran, Prashit Parikh, Jan Abendroth, Tracy L. Arakaki, Matthew C. Clifton, Thomas E. Edwards, Donald D. Lorimer, Stephen Mayclin, Bart L. Staker, Peter Myler, Krystle J. McLaughlin
The rise in antimicrobial resistance is a global health crisis and necessitates the development of novel strategies to treat infections. For example, in 2022 tuberculosis (TB) was the second leading infectious killer after COVID-19, with multi-drug-resistant strains of TB having an ∼40% fatality rate. Targeting essential biosynthetic pathways in pathogens has proven to be successful for the development of novel antimicrobial treatments. Fatty-acid synthesis (FAS) in bacteria proceeds via the type II pathway, which is substantially different from the type I pathway utilized in animals. This makes bacterial fatty-acid biosynthesis (Fab) enzymes appealing as drug targets. FabG is an essential FASII enzyme, and some bacteria, such as Mycobacterium tuberculosis, the causative agent of TB, harbor multiple homologs. FabG4 is a conserved, high-molecular-weight FabG (HMwFabG) that was first identified in M. tuberculosis and is distinct from the canonical low-molecular-weight FabG. Here, structural and functional analyses of Mycolicibacterium smegmatis FabG4, the third HMwFabG studied to date, are reported. Crystal structures of NAD+ and apo MsFabG4, along with kinetic analyses, show that MsFabG4 preferentially binds and uses NADH when reducing CoA substrates. As M. smegmatis is often used as a model organism for M. tuberculosis, these studies may aid the development of drugs to treat TB and add to the growing body of research that distinguish HMwFabGs from the archetypal low-molecular-weight FabG.
{"title":"Structural and functional characterization of FabG4 from Mycolicibacterium smegmatis","authors":"Xinping Ran, Prashit Parikh, Jan Abendroth, Tracy L. Arakaki, Matthew C. Clifton, Thomas E. Edwards, Donald D. Lorimer, Stephen Mayclin, Bart L. Staker, Peter Myler, Krystle J. McLaughlin","doi":"10.1107/S2053230X2400356X","DOIUrl":"https://doi.org/10.1107/S2053230X2400356X","url":null,"abstract":"<p>The rise in antimicrobial resistance is a global health crisis and necessitates the development of novel strategies to treat infections. For example, in 2022 tuberculosis (TB) was the second leading infectious killer after COVID-19, with multi-drug-resistant strains of TB having an ∼40% fatality rate. Targeting essential biosynthetic pathways in pathogens has proven to be successful for the development of novel antimicrobial treatments. Fatty-acid synthesis (FAS) in bacteria proceeds via the type II pathway, which is substantially different from the type I pathway utilized in animals. This makes bacterial fatty-acid biosynthesis (Fab) enzymes appealing as drug targets. FabG is an essential FASII enzyme, and some bacteria, such as <i>Mycobacterium tuberculosis</i>, the causative agent of TB, harbor multiple homologs. FabG4 is a conserved, high-molecular-weight FabG (HMwFabG) that was first identified in <i>M. tuberculosis</i> and is distinct from the canonical low-molecular-weight FabG. Here, structural and functional analyses of <i>Mycolicibacterium smegmatis</i> FabG4, the third HMwFabG studied to date, are reported. Crystal structures of NAD<sup>+</sup> and apo <i>Ms</i>FabG4, along with kinetic analyses, show that <i>Ms</i>FabG4 preferentially binds and uses NADH when reducing CoA substrates. As <i>M. smegmatis</i> is often used as a model organism for <i>M. tuberculosis</i>, these studies may aid the development of drugs to treat TB and add to the growing body of research that distinguish HMwFabGs from the archetypal low-molecular-weight FabG.</p>","PeriodicalId":7029,"journal":{"name":"Acta crystallographica. Section F, Structural biology communications","volume":null,"pages":null},"PeriodicalIF":0.9,"publicationDate":"2024-04-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140814303","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-04-30DOI: 10.1107/S2053230X24002553
Liguo Wang, Christina M. Zimanyi
High-resolution structures of biomolecules can be obtained using single-particle cryo-electron microscopy (SPA cryo-EM), and the rapidly growing number of structures solved by this method is encouraging more researchers to utilize this technique. As with other structural biology methods, sample preparation for an SPA cryo-EM data collection requires some expertise and an understanding of the strengths and limitations of the technique in order to make sensible decisions in the sample-preparation process. In this article, common strategies and pitfalls are described and practical advice is given to increase the chances of success when starting an SPA cryo-EM project.
{"title":"Cryo-EM sample preparation for high-resolution structure studies","authors":"Liguo Wang, Christina M. Zimanyi","doi":"10.1107/S2053230X24002553","DOIUrl":"10.1107/S2053230X24002553","url":null,"abstract":"<p>High-resolution structures of biomolecules can be obtained using single-particle cryo-electron microscopy (SPA cryo-EM), and the rapidly growing number of structures solved by this method is encouraging more researchers to utilize this technique. As with other structural biology methods, sample preparation for an SPA cryo-EM data collection requires some expertise and an understanding of the strengths and limitations of the technique in order to make sensible decisions in the sample-preparation process. In this article, common strategies and pitfalls are described and practical advice is given to increase the chances of success when starting an SPA cryo-EM project.</p>","PeriodicalId":7029,"journal":{"name":"Acta crystallographica. Section F, Structural biology communications","volume":null,"pages":null},"PeriodicalIF":0.9,"publicationDate":"2024-04-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140292470","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-04-30DOI: 10.1107/S2053230X24003339
Maria Cristina Nonato, Mark J. van Raaij, Jon Agirre
To find out what lies behind the articles published in Acta Cryst. F – Structural Biology Communications the journal now publishes interviews with its authors.
{"title":"Beyond publishing: introducing Interviews with authors","authors":"Maria Cristina Nonato, Mark J. van Raaij, Jon Agirre","doi":"10.1107/S2053230X24003339","DOIUrl":"10.1107/S2053230X24003339","url":null,"abstract":"<p>To find out what lies behind the articles published in <i>Acta Cryst. F – Structural Biology Communications</i> the journal now publishes interviews with its authors.</p>","PeriodicalId":7029,"journal":{"name":"Acta crystallographica. Section F, Structural biology communications","volume":null,"pages":null},"PeriodicalIF":0.9,"publicationDate":"2024-04-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140763979","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-03-04DOI: 10.1107/S2053230X24002024
Mark J. van Raaij
The current situation of scientific manuscript peer review is discussed, both generally and as applied to Acta Crystallographica F – Biological Research Communications.
本文讨论了科学手稿同行评审的现状,既有一般性的,也有适用于《晶体学报》--《生物研究通讯》的。
{"title":"The unbearable burden of peer review?","authors":"Mark J. van Raaij","doi":"10.1107/S2053230X24002024","DOIUrl":"10.1107/S2053230X24002024","url":null,"abstract":"<p>The current situation of scientific manuscript peer review is discussed, both generally and as applied to A<i>cta Crystallographica F – Biological Research Communications</i>.</p>","PeriodicalId":7029,"journal":{"name":"Acta crystallographica. Section F, Structural biology communications","volume":null,"pages":null},"PeriodicalIF":0.9,"publicationDate":"2024-03-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140020716","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-03-04DOI: 10.1107/S2053230X24000943
Sunho Hong, Jungwoo Choe
KHNYN is composed of an N-terminal KH-like RNA-binding domain and a C-terminal PIN/NYN endoribonuclease domain. It forms a complex with zinc-finger antiviral protein (ZAP), leading to the degradation of viral or cellular RNAs depending on the ZAP isoform. Here, the production, crystallization and biochemical analysis of the NYN domain (residues 477–636) of human KHNYN are presented. The NYN domain was crystallized with a heptameric single-stranded RNA from the AU-rich elements of the 3′-UTR of interferon lambda 3. The crystal belonged to space group P4132, with unit-cell parameters a = b = c = 111.3 Å, and diffacted to 1.72 Å resolution. The RNase activity of the NYN domain was demonstrated using different single-stranded RNAs, together with the binding between the NYN domain of KHNYN and the zinc-finger domain of ZAP.
KHNYN 由一个 N 端 KH 样 RNA 结合结构域和一个 C 端 PIN/NYN 内切酶结构域组成。它与锌指抗病毒蛋白(ZAP)形成复合物,根据 ZAP 异构体的不同导致病毒或细胞 RNA 的降解。本文介绍了人类 KHNYN 的 NYN 结构域(残基 477-636)的制作、结晶和生化分析。该 NYN 结构域与来自干扰素 lambda 3 的 3'-UTR 中富含 AU 元素的七聚单链 RNA 一起结晶。晶体属于空间群 P4132,单位晶胞参数为 a = b = c = 111.3 Å,衍射分辨率为 1.72 Å。利用不同的单链 RNA 验证了 NYN 结构域的 RNase 活性,以及 KHNYN 的 NYN 结构域与 ZAP 的锌指结构域之间的结合。
{"title":"Crystallization and biochemical studies of the NYN domain of human KHNYN","authors":"Sunho Hong, Jungwoo Choe","doi":"10.1107/S2053230X24000943","DOIUrl":"10.1107/S2053230X24000943","url":null,"abstract":"<p>KHNYN is composed of an N-terminal KH-like RNA-binding domain and a C-terminal PIN/NYN endoribonuclease domain. It forms a complex with zinc-finger antiviral protein (ZAP), leading to the degradation of viral or cellular RNAs depending on the ZAP isoform. Here, the production, crystallization and biochemical analysis of the NYN domain (residues 477–636) of human KHNYN are presented. The NYN domain was crystallized with a heptameric single-stranded RNA from the AU-rich elements of the 3′-UTR of interferon lambda 3. The crystal belonged to space group <i>P</i>4<sub>1</sub>32, with unit-cell parameters <i>a</i> = <i>b</i> = <i>c</i> = 111.3 Å, and diffacted to 1.72 Å resolution. The RNase activity of the NYN domain was demonstrated using different single-stranded RNAs, together with the binding between the NYN domain of KHNYN and the zinc-finger domain of ZAP.</p>","PeriodicalId":7029,"journal":{"name":"Acta crystallographica. Section F, Structural biology communications","volume":null,"pages":null},"PeriodicalIF":0.9,"publicationDate":"2024-03-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139904725","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}