scholarly journals Structural basis of membrane recognition of Toxoplasma gondii vacuole by Irgb6

2021 ◽  
Vol 5 (1) ◽  
pp. e202101149
Author(s):  
Yumiko Saijo-Hamano ◽  
Aalaa Alrahman Sherif ◽  
Ariel Pradipta ◽  
Miwa Sasai ◽  
Naoki Sakai ◽  
...  
Keyword(s):  
Toxoplasma Gondii ◽  
Conformational Changes ◽  
Molecular Mechanisms ◽  
Structural Information ◽  
Parasitophorous Vacuole ◽  
Membrane Binding ◽  
Structural Basis ◽  
In Silico Docking ◽  
Binding Interface ◽  
Dimerization Interface

The p47 immunity-related GTPase (IRG) Irgb6 plays a pioneering role in host defense against Toxoplasma gondii infection. Irgb6 is recruited to the parasitophorous vacuole membrane (PVM) formed by T. gondii and disrupts it. Despite the importance of this process, the molecular mechanisms accounting for PVM recognition by Irgb6 remain elusive because of lack of structural information on Irgb6. Here we report the crystal structures of mouse Irgb6 in the GTP-bound and nucleotide-free forms. Irgb6 exhibits a similar overall architecture to other IRGs in which GTP binding induces conformational changes in both the dimerization interface and the membrane-binding interface. The membrane-binding interface of Irgb6 assumes a unique conformation, composed of N- and C-terminal helical regions forming a phospholipid binding site. In silico docking of phospholipids further revealed membrane-binding residues that were validated through mutagenesis and cell-based assays. Collectively, these data demonstrate a novel structural basis for Irgb6 to recognize T. gondii PVM in a manner distinct from other IRGs.

scholarly journals Structural basis of membrane recognition of Toxoplasma gondii vacuole by Irgb6

2021 ◽  
Author(s):  
Yumiko Saijo Hamano ◽  
Aalaa Alrahman Sherif ◽  
Ariel Pradipta ◽  
Miwa Sasai ◽  
Naoki Sakai ◽  
...  
Keyword(s):  
Toxoplasma Gondii ◽  
Conformational Changes ◽  
Molecular Mechanisms ◽  
Structural Information ◽  
Parasitophorous Vacuole ◽  
Membrane Binding ◽  
Structural Basis ◽  
In Silico Docking ◽  
Binding Interface ◽  
Toxoplasma Gondii Infection

The p47 immunity-related GTPase (IRG) Irgb6 plays a pioneering role in host defense against Toxoplasma gondii infection. It is recruited to the parasitophorous vacuole membrane (PVM) formed by T. gondii and disrupts it. Despite the importance of this process, the molecular mechanisms accounting for PVM recognition by Irgb6 remain elusive due to lack of structural information on Irgb6. Here we report the crystal structures of mouse Irgb6 in the GTP-bound and nucleotide-free forms. Irgb6 exhibits a similar overall architecture to other IRGs in which GTP-binding induces conformational changes in both the dimerization interface and the membrane-binding interface. The membrane-binding interface of Irgb6 assumes a unique conformation, composed of N- and C-terminal helical regions forming a phospholipid binding site. In silico docking of phospholipids further revealed membrane binding residues that were validated through mutagenesis and cell-based assays. Collectively, these data demonstrate a novel structural basis for Irgb6 to recognize T. gondii PVM in a manner distinct from other IRGs.

scholarly journals Structural basis underlying complex assembly and conformational transition of the type I R-M system

2017 ◽  
Vol 114 (42) ◽  
pp. 11151-11156 ◽  
Author(s):  
Yan-Ping Liu ◽  
Qun Tang ◽  
Jie-Zhong Zhang ◽  
Li-Fei Tian ◽  
Pu Gao ◽  
...  
Keyword(s):  
Conformational Changes ◽  
Molecular Mechanisms ◽  
Conformational Transition ◽  
Structural Information ◽  
Structural Basis ◽  
Type I ◽  
Structural Comparison ◽  
Complex Assembly ◽  
Adenosyl Methionine ◽  
Restriction Modification

Type I restriction-modification (R-M) systems are multisubunit enzymes with separate DNA-recognition (S), methylation (M), and restriction (R) subunits. Despite extensive studies spanning five decades, the detailed molecular mechanisms underlying subunit assembly and conformational transition are still unclear due to the lack of high-resolution structural information. Here, we report the atomic structure of a type I MTase complex (2M+1S) bound to DNA and cofactor S-adenosyl methionine in the “open” form. The intermolecular interactions between M and S subunits are mediated by a four-helix bundle motif, which also determines the specificity of the interaction. Structural comparison between open and previously reported low-resolution “closed” structures identifies the huge conformational changes within the MTase complex. Furthermore, biochemical results show that R subunits prefer to load onto the closed form MTase. Based on our results, we proposed an updated model for the complex assembly. The work reported here provides guidelines for future applications in molecular biology.

scholarly journals Structural basis of SARS-CoV-2 spike protein induced by ACE2

2020 ◽  
Author(s):  
Tomer Meirson ◽  
David Bomze ◽  
Gal Markel
Keyword(s):  
Conformational Changes ◽  
Viral Entry ◽  
Structural Information ◽  
Active Form ◽  
Intramolecular Interactions ◽  
Host Cells ◽  
Structural Basis ◽  
S Protein ◽  
Binding Interface ◽  
Recent Emergence

AbstractMotivationThe recent emergence of the novel SARS-coronavirus 2 (SARS-CoV-2) and its international spread pose a global health emergency. The viral spike (S) glycoprotein binds the receptor (angiotensin-converting enzyme 2) ACE2 and promotes SARS-CoV-2 entry into host cells. The trimeric S protein binds the receptor using the distal receptor-binding domain (RBD) causing conformational changes in S protein that allow priming by host cell proteases. Unravelling the dynamic structural features used by SARS-CoV-2 for entry might provide insights into viral transmission and reveal novel therapeutic targets. Using structures determined by X-ray crystallography and cryo-EM, we performed structural analysis and atomic comparisons of the different conformational states adopted by the SARS-CoV-2-RBD.ResultsHere, we determined the key structural components induced by the receptor and characterized their intramolecular interactions. We show that κ-helix (also known as polyproline II) is a predominant structure in the binding interface and in facilitating the conversion to the active form of the S protein. We demonstrate a series of conversions between switch-like κ-helix and β-strand, and conformational variations in a set of short α-helices which affect the proximal hinge region. This conformational changes lead to an alternating pattern in conserved disulfide bond configurations positioned at the hinge, indicating a possible disulfide exchange, an important allosteric switch implicated in viral entry of various viruses, including HIV and murine coronavirus. The structural information presented herein enables us to inspect and understand the important dynamic features of SARS-CoV-2-RBD and propose a novel potential therapeutic strategy to block viral entry. Overall, this study provides guidance for the design and optimization of structure-based intervention strategies that target SARS-CoV-2.

Crystal structures of Magnaporthe oryzae trehalose-6-phosphate synthase (MoTps1) suggest a model for catalytic process of Tps1

2019 ◽  
Vol 476 (21) ◽  
pp. 3227-3240 ◽  
Author(s):  
Shanshan Wang ◽  
Yanxiang Zhao ◽  
Long Yi ◽  
Minghe Shen ◽  
Chao Wang ◽  
...  
Keyword(s):  
Crystal Structures ◽  
Conformational Changes ◽  
Magnaporthe Oryzae ◽  
Structural Information ◽  
Complex Structure ◽  
Critical Role ◽  
Catalytic Process ◽  
Structural Basis ◽  
Salt Bridges ◽  
Terminal Domain

Trehalose-6-phosphate (T6P) synthase (Tps1) catalyzes the formation of T6P from UDP-glucose (UDPG) (or GDPG, etc.) and glucose-6-phosphate (G6P), and structural basis of this process has not been well studied. MoTps1 (Magnaporthe oryzae Tps1) plays a critical role in carbon and nitrogen metabolism, but its structural information is unknown. Here we present the crystal structures of MoTps1 apo, binary (with UDPG) and ternary (with UDPG/G6P or UDP/T6P) complexes. MoTps1 consists of two modified Rossmann-fold domains and a catalytic center in-between. Unlike Escherichia coli OtsA (EcOtsA, the Tps1 of E. coli), MoTps1 exists as a mixture of monomer, dimer, and oligomer in solution. Inter-chain salt bridges, which are not fully conserved in EcOtsA, play primary roles in MoTps1 oligomerization. Binding of UDPG by MoTps1 C-terminal domain modifies the substrate pocket of MoTps1. In the MoTps1 ternary complex structure, UDP and T6P, the products of UDPG and G6P, are detected, and substantial conformational rearrangements of N-terminal domain, including structural reshuffling (β3–β4 loop to α0 helix) and movement of a ‘shift region' towards the catalytic centre, are observed. These conformational changes render MoTps1 to a ‘closed' state compared with its ‘open' state in apo or UDPG complex structures. By solving the EcOtsA apo structure, we confirmed that similar ligand binding induced conformational changes also exist in EcOtsA, although no structural reshuffling involved. Based on our research and previous studies, we present a model for the catalytic process of Tps1. Our research provides novel information on MoTps1, Tps1 family, and structure-based antifungal drug design.

scholarly journals Plasma membrane association facilitates conformational changes in the Marburg virus protein VP40 dimer

RSC Advances ◽  
2017 ◽  
Vol 7 (37) ◽  
pp. 22741-22748 ◽  
Author(s):  
Nisha Bhattarai ◽  
Jeevan B. GC ◽  
Bernard S. Gerstman ◽  
Robert V. Stahelin ◽  
Prem P. Chapagain
Keyword(s):  
Plasma Membrane ◽  
Conformational Changes ◽  
Ebola Virus ◽  
Membrane Binding ◽  
Marburg Virus ◽  
Virus Protein ◽  
Membrane Association ◽  
Binding Interface

The membrane binding interface of the Marburg virus protein mVP40 dimer differs from that of the Ebola virus eVP40 dimer but membrane binding allows conformational changes in mVP40 that makes it structurally similar to the eVP40 dimer.

scholarly journals Structure and inhibitor binding characterization of oncogenic MLLT1 mutants

2021 ◽  
Author(s):  
Xiaomin Ni ◽  
Allyn T. Londregan ◽  
Dafydd R. Owen ◽  
Stefan Knapp ◽  
Apirat Chaikuad
Keyword(s):  
Conformational Changes ◽  
Structural Basis ◽  
Wild Type ◽  
Structural Comparison ◽  
Inhibitor Binding ◽  
Binding Interface ◽  
Association Behavior ◽  
Self Association ◽  
Potential Strategy

AbstractDysfunction of YEATS-domain-containing MLLT1, an acetyl/acyl-lysine dependent epigenetic reader domain, has been implicated in the development of aggressive cancers. Mutations in the YEATS domain have been recently reported as a cause of MLLT1 aberrant reader function. However, structural basis for the reported alterations in affinity for acetyled/acylated histone has remained elusive. Here, we report the crystal structures of both insertion and substitution present in cancer, revealing significant conformational changes of the YEATS-domain loop 8. Structural comparison demonstrates that such alteration not only altered the binding interface for acetylated/acylated histones, but the sequence alterations in the T1 loop may enable dimeric assembly consistent inducing self-association behavior. Nevertheless, we show that also the MLLT1 mutants can be targeted by developed acetyllysine mimetic inhibitors with affinities similarly to wild type. Our report provides a structural basis for the altered behaviors and potential strategy for targeting oncogenic MLLT1 mutants.

scholarly journals Determinants shaping the nanoscale architecture of the mouse rod outer segment

2021 ◽  
Author(s):  
Matthias Pöge ◽  
Julia Mahamid ◽  
Sanae S Imanishi ◽  
Jürgen M Plitzko ◽  
Krzysztof Palczewski ◽  
...  
Keyword(s):  
Outer Segment ◽  
Molecular Mechanisms ◽  
Structural Information ◽  
Structural Basis ◽  
Radius Of Curvature ◽  
Rod Photoreceptor ◽  
Molecular Architecture ◽  
Structural Determinants ◽  
Rod Outer Segment ◽  
Wide Range

The unique membrane organization of the rod outer segment (ROS), the specialized sensory cilium of rod photoreceptor cells, provides the foundation for phototransduction, the initial step in vision. ROS architecture is characterized by a stack of identically shaped and tightly packed membrane disks loaded with the visual receptor rhodopsin. A wide range of genetic aberrations compromise ROS ultrastructure, impairing photoreceptor viability and function. Yet, the structural basis giving rise to the remarkable long range order of ROS membrane stacks and the molecular mechanisms underlying genetically inherited diseases remain elusive. Here, cryo-electron tomography (cryo-ET) performed on native ROS at molecular resolution provides insights into key structural determinants of ROS membrane architecture. Our data reveal the existence of two molecular connectors/spacers which likely contribute to the nanometer scale precise stacking of the ROS disks. We further show that the extreme radius of curvature at the disk rims is enforced by a continuous supramolecular assembly composed of peripherin-2 (PRPH2) and rod outer segment membrane protein 1 (ROM1) tetramers. We suggest that, together these molecular assemblies constitute the structural basis of the highly specialized ROS functional architecture. Cryo-ET therefore provides novel quantitative and structural information on the molecular architecture in ROS and insights into possible mechanisms underlying pathologies of certain PRPH2 mutations leading to blindness.

scholarly journals Structural basis of Toxoplasma gondii Perforin-Like Protein 1 membrane interaction and activity during egress

2018 ◽  
Author(s):  
Alfredo J. Guerra ◽  
Ou Zhang ◽  
Constance M. E. Bahr ◽  
My-Hang Huynh ◽  
James DelProposto ◽  
...  
Keyword(s):  
Toxoplasma Gondii ◽  
Host Cell ◽  
Membrane Binding ◽  
Membrane Attack Complex ◽  
Direct Repeat ◽  
Tryptophan Fluorescence ◽  
Cytolytic Activity ◽  
Complex Life Cycle ◽  
Structural Basis ◽  
Terminal Domain

AbstractIntracellular pathogens must egress from the host cell to continue their infectious cycle. Apicomplexans are a phylum of intracellular protozoans that have evolved members of the membrane attack complex and perforin (MACPF) family of pore forming proteins to disrupt cellular membranes for traversing cells during tissue migration or egress from a replicative vacuole following intracellular reproduction. Previous work showed that the apicomplexan Toxoplasma gondii secretes a perforin-like protein (TgPLP1) that contains a C-terminal Domain (CTD) which is necessary for efficient parasite egress. However, the structural basis for CTD membrane binding and egress competency remained unknown. Here, we present evidence that TgPLP1 CTD prefers binding lipids that are abundant in the inner leaflet of the lipid bilayer. Additionally, solving the high-resolution crystal structure of the TgPLP1 APCβ domain within the CTD reveals an unusual double-layered β-prism fold that resembles only one other protein of known structure. Three direct repeat sequences comprise subdomains, with each constituting a wall of the β-prism fold. One subdomain features a protruding hydrophobic loop with an exposed tryptophan at its tip. Spectrophotometric measurements of intrinsic tryptophan fluorescence are consistent with insertion of the hydrophobic loop into a target membrane. Using CRISPR/Cas9 gene editing we show that parasite strains bearing mutations in the hydrophobic loop, including alanine substitution of the tip tryptophan, are equally deficient in egress as a strain lacking TgPLP1 altogether. Taken together our findings suggest a crucial role for the hydrophobic loop in anchoring TgPLP1 to the membrane to support its cytolytic activity and egress function.Author SummaryToxoplasma gondii has a complex life cycle that involves active invasion of the host cell, the formation of a replicative compartment, and egress from the replicative niche. T. gondii encodes a pore-forming protein, TgPLP1, that contains a C-terminal domain that is crucial for efficient exit from both the parasite containing vacuole and the host cell. However, the mechanism by which TgPLP1 recognizes and binds to the appropriate membrane is unclear. Here we use a combination of biochemistry, structural biology, and parasitology to identify the a preference of TgPLP1 for specific lipids and show that a loop within the structure of the C-terminal domain inserts into the membrane and is necessary for egress from the parasite containing vacuole. Our study sheds light into the determinants of membrane binding in TgPLP1 which may inform the overall mechanism of pore formation in similar systems

Redox- and pH-linked conformational changes in triheme cytochrome PpcA from Geobacter sulfurreducens

2017 ◽  
Vol 474 (2) ◽  
pp. 231-246 ◽  
Author(s):  
Leonor Morgado ◽  
Marta Bruix ◽  
P. Raj Pokkuluri ◽  
Carlos A. Salgueiro ◽  
David L. Turner
Keyword(s):  
Conformational Changes ◽  
Solution Structure ◽  
Molecular Mechanisms ◽  
Three Dimensional ◽  
Axial Ligand ◽  
Geobacter Sulfurreducens ◽  
Cellular Growth ◽  
Dimensional Structure ◽  
Structural Basis ◽  
Natural Habitats

The periplasmic triheme cytochrome PpcA from Geobacter sulfurreducens is highly abundant; it is the likely reservoir of electrons to the outer surface to assist the reduction of extracellular terminal acceptors; these include insoluble metal oxides in natural habitats and electrode surfaces from which electricity can be harvested. A detailed thermodynamic characterization of PpcA showed that it has an important redox-Bohr effect that might implicate the protein in e−/H+ coupling mechanisms to sustain cellular growth. This functional mechanism requires control of both the redox state and the protonation state. In the present study, isotope-labeled PpcA was produced and the three-dimensional structure of PpcA in the oxidized form was determined by NMR. This is the first solution structure of a G. sulfurreducens cytochrome in the oxidized state. The comparison of oxidized and reduced structures revealed that the heme I axial ligand geometry changed and there were other significant changes in the segments near heme I. The pH-linked conformational rearrangements observed in the vicinity of the redox-Bohr center, both in the oxidized and reduced structures, constitute the structural basis for the differences observed in the pKa values of the redox-Bohr center, providing insights into the e−/H+ coupling molecular mechanisms driven by PpcA in G. sulfurreducens.

scholarly journals Determinants shaping the nanoscale architecture of the mouse rod outer segment

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Matthias Pöge ◽  
Julia Mahamid ◽  
Sanae S Imanishi ◽  
Jürgen M Plitzko ◽  
Krzysztof Palczewski ◽  
...  
Keyword(s):  
Outer Segment ◽  
Molecular Mechanisms ◽  
Structural Information ◽  
Structural Basis ◽  
Radius Of Curvature ◽  
Rod Photoreceptor ◽  
Molecular Architecture ◽  
Structural Determinants ◽  
Rod Outer Segment ◽  
Wide Range

The unique membrane organization of the rod outer segment (ROS), the specialized sensory cilium of rod photoreceptor cells, provides the foundation for phototransduction, the initial step in vision. ROS architecture is characterized by a stack of identically shaped and tightly packed membrane disks loaded with the visual receptor rhodopsin. A wide range of genetic aberrations have been reported to compromise ROS ultrastructure, impairing photoreceptor viability and function. Yet, the structural basis giving rise to the remarkably precise arrangement of ROS membrane stacks and the molecular mechanisms underlying genetically inherited diseases remain elusive. Here, cryo-electron tomography (cryo-ET) performed on native ROS at molecular resolution provides insights into key structural determinants of ROS membrane architecture. Our data confirm the existence of two previously observed molecular connectors/spacers which likely contribute to the nanometer-scale precise stacking of the ROS disks. We further provide evidence that the extreme radius of curvature at the disk rims is enforced by a continuous supramolecular assembly composed of peripherin-2 (PRPH2) and rod outer segment membrane protein 1 (ROM1) oligomers. We suggest that together these molecular assemblies constitute the structural basis of the highly specialized ROS functional architecture. Our Cryo-ET data provide novel quantitative and structural information on the molecular architecture in ROS and substantiate previous results on proposed mechanisms underlying pathologies of certain PRPH2 mutations leading to blindness.

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