Elucidating the Structural Basis of Protein Domain Coupling in Spectrin Repeat Domains

Recent epidemics demonstrate the global threat of Zika virus (ZIKV), a flavivirus transmitted by mosquitoes. Although infection is usually asymptomatic or mild, newborns of infected mothers can display severe symptoms, including neurodevelopmental abnormalities and microcephaly. Given the large-scale spread, symptom severity, and lack of treatment or prophylaxis, a safe and effective ZIKV vaccine is urgently needed. However, vaccine design is complicated by concern that elicited antibodies (Abs) may cross-react with other flaviviruses that share a similar envelope protein, such as dengue virus, West Nile virus, and yellow fever virus. This cross-reactivity may worsen symptoms of a subsequent infection through Ab-dependent enhancement. To better understand the neutralizing Ab response and risk of Ab-dependent enhancement, further information on germline Ab binding to ZIKV and the maturation process that gives rise to potently neutralizing Abs is needed. Here we use binding and structural studies to compare mature and inferred-germline Ab binding to envelope protein domain III of ZIKV and other flaviviruses. We show that affinity maturation of the light-chain variable domain is important for strong binding of the recurrent VH3-23/VK1-5 neutralizing Abs to ZIKV envelope protein domain III, and identify interacting residues that contribute to weak, cross-reactive binding to West Nile virus. These findings provide insight into the affinity maturation process and potential cross-reactivity of VH3-23/VK1-5 neutralizing Abs, informing precautions for protein-based vaccines designed to elicit germline versions of neutralizing Abs.

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The structure of a plant-specific partitivirus capsid reveals a unique coat protein domain architecture with an intrinsically disordered protrusion

Communications Biology ◽

10.1038/s42003-021-02687-w ◽

2021 ◽

Vol 4 (1) ◽

Author(s):

Matthew Byrne ◽

Aseem Kashyap ◽

Lygie Esquirol ◽

Neil Ranson ◽

Frank Sainsbury

Keyword(s):

Coat Protein ◽

Domain Architecture ◽

Plant Viruses ◽

Protein Domain ◽

Wild Plants ◽

Structural Basis ◽

Coat Proteins ◽

Intrinsically Disordered ◽

Binding Interface ◽

Protein Dimers

AbstractPersistent plant viruses may be the most common viruses in wild plants. A growing body of evidence for mutualism between such viruses and their hosts, suggests that they play an important role in ecology and agriculture. Here we present the capsid structure of a plant-specific partitivirus, Pepper cryptic virus 1, at 2.9 Å resolution by Cryo-EM. Structural features, including the T = 1 arrangement of 60 coat protein dimers, are shared with fungal partitiviruses and the picobirnavirus lineage of dsRNA viruses. However, the topology of the capsid is markedly different with protrusions emanating from, and partly comprising, the binding interface of coat protein dimers. We show that a disordered region at the apex of the protrusion is not required for capsid assembly and represents a hypervariable site unique to, and characteristic of, the plant-specific partitiviruses. These results suggest a structural basis for the acquisition of additional functions by partitivirus coat proteins that enables mutualistic relationships with diverse plant hosts.

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Structural basis underlying strong interactions between ankyrins and spectrins

10.1101/2020.03.04.976142 ◽

2020 ◽

Author(s):

Jianchao Li ◽

Keyu Chen ◽

Ruichi Zhu ◽

Mingjie Zhang

Keyword(s):

Plasma Membranes ◽

Molecular Mechanisms ◽

Insertion Site ◽

Structural Basis ◽

Membrane Microdomains ◽

Strong Interactions ◽

Structural Investigations ◽

Spectrin Repeat ◽

Subcellular Membrane ◽

Junction Site

AbstractAnkyrins (encoded by ANK1/2/3 corresponding to Ankyrin-R/B/G or AnkR/B/G), via binding to spectrins, connect plasma membranes with actin cytoskeleton to maintain mechanical strengths and to modulate excitabilities of diverse cells such as neurons, muscle cells, and erythrocytes. Cellular and genetic evidences suggest that each isoform of ankyrins pairs with a specific β-spectrin in discrete subcellular membrane microdomains for distinct functions, though the molecular mechanisms underlying such ankyrin/β-spectrin pairings are unknown. In this study, we discover that a conserved and short extension N-terminal to the ZU5N-ZU5C-UPA tandem (exZZU) is critical for each ankyrin to bind to β-spectrins with high affinities. Structures of AnkB/G exZZU in complex with spectrin repeats13-15 of β2/β4-spectrins solved here reveal that the extension sequence of exZZU forms an additional β-strand contributing to the structural stability and enhanced affinity of each ZU5N/spectrin repeat interaction. The junction site between the extension and ZU5N is exactly the position of a splicing-mediated miniexon insertion site of AnkB/G. The complex structures further reveal that the UPA domain of exZZU directly participates in spectrin binding. Formation of the exZZU supramodule juxtaposes the ZU5N and UPA domains for simultaneous interacting with spectrin repeats 14 and 15. However, our biochemical and structural investigations indicate that the direct and strong interactions between ankyrins and β-spectrins do not appear to determine their pairing specificities. Therefore, there likely exists additional mechanism(s) for modulating functional pairings between ankyrins and β-spectrins in cells.

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Structural basis for activation and non-canonical catalysis of the Rap GTPase activating protein domain of plexin

eLife ◽

10.7554/elife.01279 ◽

2013 ◽

Vol 2 ◽

Cited By ~ 46

Author(s):

Yuxiao Wang ◽

Heath G Pascoe ◽

Chad A Brautigam ◽

Huawei He ◽

Xuewu Zhang

Keyword(s):

Cell Surface ◽

Axon Guidance ◽

Active Site ◽

Large Scale ◽

Catalytic Mechanism ◽

Protein Domain ◽

Structural Basis ◽

Gtpase Activating Protein ◽

Gtp Hydrolysis ◽

Surface Receptors

Plexins are cell surface receptors that bind semaphorins and transduce signals for regulating neuronal axon guidance and other processes. Plexin signaling depends on their cytoplasmic GTPase activating protein (GAP) domain, which specifically inactivates the Ras homolog Rap through an ill-defined non-canonical catalytic mechanism. The plexin GAP is activated by semaphorin-induced dimerization, the structural basis for which remained unknown. Here we present the crystal structures of the active dimer of zebrafish PlexinC1 cytoplasmic region in the apo state and in complex with Rap. The structures show that the dimerization induces a large-scale conformational change in plexin, which opens the GAP active site to allow Rap binding. Plexin stabilizes the switch II region of Rap in an unprecedented conformation, bringing Gln63 in Rap into the active site for catalyzing GTP hydrolysis. The structures also explain the unique Rap-specificity of plexins. Mutational analyses support that these mechanisms underlie plexin activation and signaling.

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Phosphopantetheinyl transferase binding and inhibition by amidino-urea and hydroxypyrimidinethione compounds

Scientific Reports ◽

10.1038/s41598-021-97197-4 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Coralie Carivenc ◽

Laurent Maveyraud ◽

Claire Blanger ◽

Stéphanie Ballereau ◽

Coralie Roy-Camille ◽

...

Keyword(s):

Pseudomonas Aeruginosa ◽

Pathogenic Bacteria ◽

Acyl Carrier Protein ◽

Mycobacterium Abscessus ◽

Protein Domain ◽

Structural Basis ◽

Inhibition Mechanism ◽

Phosphopantetheinyl Transferase ◽

Phosphopantetheinyl Transferases

AbstractOwing to their role in activating enzymes essential for bacterial viability and pathogenicity, phosphopantetheinyl transferases represent novel and attractive drug targets. In this work, we examined the inhibitory effect of the aminido-urea 8918 compound against the phosphopantetheinyl transferases PptAb from Mycobacterium abscessus and PcpS from Pseudomonas aeruginosa, two pathogenic bacteria associated with cystic fibrosis and bronchiectasis, respectively. Compound 8918 exhibits inhibitory activity against PptAb but displays no activity against PcpS in vitro, while no antimicrobial activity against Mycobacterium abscessus or Pseudomonas aeruginosa could be detected. X-ray crystallographic analysis of 8918 bound to PptAb-CoA alone and in complex with an acyl carrier protein domain in addition to the crystal structure of PcpS in complex with CoA revealed the structural basis for the inhibition mechanism of PptAb by 8918 and its ineffectiveness against PcpS. Finally, in crystallo screening of potent inhibitors from the National Cancer Institute library identified a hydroxypyrimidinethione derivative that binds PptAb. Both compounds could serve as scaffolds for the future development of phosphopantetheinyl transferases inhibitors.

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Decision letter: Structural basis for activation and non-canonical catalysis of the Rap GTPase activating protein domain of plexin

10.7554/elife.01279.019 ◽

2013 ◽

Keyword(s):

Protein Domain ◽

Structural Basis ◽

Gtpase Activating Protein

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Author response: Structural basis for activation and non-canonical catalysis of the Rap GTPase activating protein domain of plexin

10.7554/elife.01279.020 ◽

2013 ◽

Cited By ~ 3

Author(s):

Yuxiao Wang ◽

Heath G Pascoe ◽

Chad A Brautigam ◽

Huawei He ◽

Xuewu Zhang

Keyword(s):

Protein Domain ◽

Author Response ◽

Structural Basis ◽

Gtpase Activating Protein

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Structures of the spectrin-ankyrin interaction binding domains

Blood ◽

10.1182/blood-2008-10-184358 ◽

2009 ◽

Vol 113 (22) ◽

pp. 5385-5393 ◽

Cited By ~ 65

Author(s):

Jonathan J. Ipsaro ◽

Lei Huang ◽

Alfonso Mondragón

Keyword(s):

Consensus Sequence ◽

Membrane Skeleton ◽

Binding Domain ◽

Structural Basis ◽

Spectrin Repeat ◽

The Core ◽

Binding Domains ◽

Novel Structure ◽

Important Region ◽

Spectrin Cytoskeleton

As key components of the erythrocyte membrane skeleton, spectrin and ankyrin specifically interact to tether the spectrin cytoskeleton to the cell membrane. The structure of the spectrin binding domain of ankyrin and the ankyrin binding domain of spectrin have been solved to elucidate the structural basis for ankyrin-spectrin recognition. The structure of repeats 14 and 15 of spectrin shows that these repeats are similar to all other spectrin repeats. One feature that could account for the preference of ankyrin for these repeats is the presence of a conserved, negatively charged patch on one side of repeat 14. The structure of the ankyrin ZU5 domain shows a novel structure containing a β core. The structure reveals that the canonical ZU5 consensus sequence is likely to be missing an important region that codes for a β strand that forms part of the core of the domain. In addition, a positively charged region is suggestive of a binding surface for the negatively charged spectrin repeat 14. Previously reported mutants of ankyrin that map to this region lie mostly on the surface of the protein, although at least one is likely to be part of the core.

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Organization of tight junctions in the choroid plexus epithelium

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100082546 ◽

1978 ◽

Vol 36 (2) ◽

pp. 336-337

Author(s):

B. Van Deurs ◽

J. K. Koehler

Keyword(s):

Choroid Plexus ◽

Present Report ◽

Freeze Fracture ◽

Barrier Properties ◽

Cell Junctions ◽

Structural Basis ◽

Apical Surface ◽

Choroid Plexus Epithelium ◽

Solid Nitrogen ◽

Special Composition

The choroid plexus epithelium constitutes a blood-cerebrospinal fluid (CSF) barrier, and is involved in regulation of the special composition of the CSF. The epithelium is provided with an ouabain-sensitive Na/K-pump located at the apical surface, actively pumping ions into the CSF. The choroid plexus epithelium has been described as “leaky” with a low transepithelial resistance, and a passive transepithelial flux following a paracellular route (intercellular spaces and cell junctions) also takes place. The present report describes the structural basis for these “barrier” properties of the choroid plexus epithelium as revealed by freeze fracture.Choroid plexus from the lateral, third and fourth ventricles of rats were used. The tissue was fixed in glutaraldehyde and stored in 30% glycerol. Freezing was performed either in liquid nitrogen-cooled Freon 22, or directly in a mixture of liquid and solid nitrogen prepared in a special vacuum chamber. The latter method was always used, and considered necessary, when preparations of complementary (double) replicas were made.

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High resolution spot scan imaging of frozen, hydrated actin bundles with 400kV electrons

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100122964 ◽

1992 ◽

Vol 50 (1) ◽

pp. 512-513

Author(s):

J. Jakana ◽

M.F. Schmid ◽

P. Matsudaira ◽

W. Chiu

Keyword(s):

High Resolution ◽

Binding Proteins ◽

Actin Binding ◽

Eukaryotic Cells ◽

Dimensional Structure ◽

Structural Basis ◽

Actin Bundle ◽

Actin Bundles ◽

3 Dimensional ◽

Macromolecular Assembly

Actin is a protein found in all eukaryotic cells. In its polymerized form, the cells use it for motility, cytokinesis and for cytoskeletal support. An example of this latter class is the actin bundle in the acrosomal process from the Limulus sperm. The different functions actin performs seem to arise from its interaction with the actin binding proteins. A 3-dimensional structure of this macromolecular assembly is essential to provide a structural basis for understanding this interaction in relationship to its development and functions.

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