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Molecular BioSystems May 2008Transient receptor potential (TRP) channels are rapidly gaining attention as important receptors and transducers of diverse sensory and environmental cues. Recent... (Review)
Review
Transient receptor potential (TRP) channels are rapidly gaining attention as important receptors and transducers of diverse sensory and environmental cues. Recent progress in the field has provided new insights into the structure and function of the ankyrin repeat motifs present in the N-terminal cytosolic domain of many TRP channels. The topics addressed in this Highlight include the structural features of canonical ankyrin repeats, new clues into the functions these repeats perform in cells, and how this information can be applied to develop further experiments on TRP channels and other proteins containing ankyrin repeats.
Topics: Ankyrin Repeat; Ligands; Models, Molecular; Protein Conformation; Transient Receptor Potential Channels
PubMed: 18414734
DOI: 10.1039/b801481g -
Frontiers in Bioinformatics 2021Protein repeats are short, highly similar peptide motifs that occur several times within a single protein, for example the TPR and Ankyrin repeats. Understanding the... (Review)
Review
Protein repeats are short, highly similar peptide motifs that occur several times within a single protein, for example the TPR and Ankyrin repeats. Understanding the role of mutation in these proteins is complicated by the competing facts that 1) the repeats are much more restricted to a set sequence than non-repeat proteins, so mutations should be harmful much more often because there are more residues that are heavily restricted due to the need of the sequence to repeat and 2) the symmetry of the repeats in allows the distribution of functional contributions over a number of residues so that sometimes no specific site is singularly responsible for function (unlike enzymatic active site catalytic residues). To address this issue, we review the effects of mutations in a number of natural repeat proteins from the tetratricopeptide and Ankyrin repeat families. We find that mutations are context dependent. Some mutations are indeed highly disruptive to the function of the protein repeats while mutations in identical positions in other repeats in the same protein have little to no effect on structure or function.
PubMed: 36303725
DOI: 10.3389/fbinf.2021.696368 -
Nature Communications Jun 2023Crosstalk between ion channels and small GTPases is critical during homeostasis and disease, but little is known about the structural underpinnings of these...
Crosstalk between ion channels and small GTPases is critical during homeostasis and disease, but little is known about the structural underpinnings of these interactions. TRPV4 is a polymodal, calcium-permeable cation channel that has emerged as a potential therapeutic target in multiple conditions. Gain-of-function mutations also cause hereditary neuromuscular disease. Here, we present cryo-EM structures of human TRPV4 in complex with RhoA in the ligand-free, antagonist-bound closed, and agonist-bound open states. These structures reveal the mechanism of ligand-dependent TRPV4 gating. Channel activation is associated with rigid-body rotation of the intracellular ankyrin repeat domain, but state-dependent interaction with membrane-anchored RhoA constrains this movement. Notably, many residues at the TRPV4-RhoA interface are mutated in disease and perturbing this interface by introducing mutations into either TRPV4 or RhoA increases TRPV4 channel activity. Together, these results suggest that RhoA serves as an auxiliary subunit for TRPV4, regulating TRPV4-mediated calcium homeostasis and disruption of TRPV4-RhoA interactions can lead to TRPV4-related neuromuscular disease. These insights will help facilitate TRPV4 therapeutics development.
Topics: Humans; Ankyrin Repeat; Calcium; Mutation; TRPV Cation Channels; rhoA GTP-Binding Protein
PubMed: 37353484
DOI: 10.1038/s41467-023-39345-0 -
PLoS Computational Biology Dec 2015Ankyrin repeat containing proteins are one of the most abundant solenoid folds. Usually implicated in specific protein-protein interactions, these proteins are readily...
Ankyrin repeat containing proteins are one of the most abundant solenoid folds. Usually implicated in specific protein-protein interactions, these proteins are readily amenable for design, with promising biotechnological and biomedical applications. Studying repeat protein families presents technical challenges due to the high sequence divergence among the repeating units. We developed and applied a systematic method to consistently identify and annotate the structural repetitions over the members of the complete Ankyrin Repeat Protein Family, with increased sensitivity over previous studies. We statistically characterized the number of repeats, the folding of the repeat-arrays, their structural variations, insertions and deletions. An energetic analysis of the local frustration patterns reveal the basic features underlying fold stability and its relation to the functional binding regions. We found a strong linear correlation between the conservation of the energetic features in the repeat arrays and their sequence variations, and discuss new insights into the organization and function of these ubiquitous proteins.
Topics: Amino Acid Sequence; Ankyrin Repeat; Ankyrins; Computer Simulation; Energy Transfer; Models, Chemical; Models, Molecular; Molecular Sequence Data; Sequence Analysis, Protein
PubMed: 26691182
DOI: 10.1371/journal.pcbi.1004659 -
Nature Structural & Molecular Biology Jul 2022The cytoskeleton of a red blood cell (RBC) is anchored to the cell membrane by the ankyrin complex. This complex is assembled during RBC genesis and comprises primarily...
The cytoskeleton of a red blood cell (RBC) is anchored to the cell membrane by the ankyrin complex. This complex is assembled during RBC genesis and comprises primarily band 3, protein 4.2 and ankyrin, whose mutations contribute to numerous human inherited diseases. High-resolution structures of the ankyrin complex have been long sought-after to understand its assembly and disease-causing mutations. Here, we analyzed native complexes on the human RBC membrane by stepwise fractionation. Cryo-electron microscopy structures of nine band-3-associated complexes reveal that protein 4.2 stabilizes the cytoplasmic domain of band 3 dimer. In turn, the superhelix-shaped ankyrin binds to this protein 4.2 via ankyrin repeats (ARs) 6-13 and to another band 3 dimer via ARs 17-20, bridging two band 3 dimers in the ankyrin complex. Integration of these structures with both prior data and our biochemical data supports a model of ankyrin complex assembly during erythropoiesis and identifies interactions essential for the mechanical stability of RBC.
Topics: Ankyrin Repeat; Ankyrins; Cryoelectron Microscopy; Cytoskeleton; Erythrocyte Membrane; Humans
PubMed: 35655099
DOI: 10.1038/s41594-022-00779-7 -
Journal of Molecular Biology Feb 2008Full-consensus designed ankyrin repeat proteins were designed with one to six identical repeats flanked by capping repeats. These proteins express well in Escherichia...
Full-consensus designed ankyrin repeat proteins were designed with one to six identical repeats flanked by capping repeats. These proteins express well in Escherichia coli as soluble monomers. Compared to our previously described designed ankyrin repeat protein library, randomized positions have now been fixed according to sequence statistics and structural considerations. Their stability increases with length and is even higher than that of library members, and those with more than three internal repeats are resistant to denaturation by boiling or guanidine hydrochloride. Full denaturation requires their heating in 5 M guanidine hydrochloride. The folding and unfolding kinetics of the proteins with up to three internal repeats were analyzed, as the other proteins could not be denatured. Folding is monophasic, with a rate that is nearly identical for all proteins ( approximately 400-800 s(-1)), indicating that essentially the same transition state must be crossed, possibly the folding of a single repeat. In contrast, the unfolding rate decreases by a factor of about 10(4) with increasing repeat number, directly reflecting thermodynamic stability in these extraordinarily slow denaturation rates. The number of unfolding phases also increases with repeat number. We analyzed the folding thermodynamics and kinetics both by classical two-state and three-state cooperative models and by an Ising-like model, where repeats are considered as two-state folding units that can be stabilized by interacting with their folded nearest neighbors. This Ising model globally describes both equilibrium and kinetic data very well and allows for a detailed explanation of the ankyrin repeat protein folding mechanism.
Topics: Amino Acid Sequence; Ankyrin Repeat; Ankyrins; Circular Dichroism; Escherichia coli; Gene Expression; Kinetics; Models, Molecular; Molecular Sequence Data; Protein Folding; Spectrometry, Fluorescence
PubMed: 18164721
DOI: 10.1016/j.jmb.2007.11.046 -
Viruses Oct 2022Designed ankyrin repeat proteins (DARPins) are engineered proteins comprising consensus designed ankyrin repeats as scaffold. Tightly packed repeats form a continuous... (Review)
Review
Designed ankyrin repeat proteins (DARPins) are engineered proteins comprising consensus designed ankyrin repeats as scaffold. Tightly packed repeats form a continuous hydrophobic core and a large groove-like solvent-accessible surface that creates a binding surface. DARPin domains recognizing a target of interest with high specificity and affinity can be generated using a synthetic combinatorial library and in vitro selection methods. They can be linked together in a single molecule to build multispecific and multifunctional proteins without affecting expression or function. The modular architecture of DARPins offers unprecedented possibilities of design and opens avenues for innovative antiviral strategies.
Topics: Designed Ankyrin Repeat Proteins; Virus Internalization; Ankyrin Repeat; Proteins; HIV Fusion Inhibitors; Solvents
PubMed: 36298797
DOI: 10.3390/v14102242 -
Nature Communications Sep 2022Over 70% of vascular flowering plants engage in endosymbiotic associations with arbuscular mycorrhizal (AM) fungi. VAPYRIN (VPY) is a plant protein that is required for...
Over 70% of vascular flowering plants engage in endosymbiotic associations with arbuscular mycorrhizal (AM) fungi. VAPYRIN (VPY) is a plant protein that is required for intracellular accommodation of AM fungi but how it functions is still unclear. VPY has a large ankyrin repeat domain with potential for interactions with multiple proteins. Here we show that overexpression of the ankyrin repeat domain results in a vpy-like phenotype, consistent with the sequestration of interacting proteins. We identify distinct ankyrin repeats that are essential for intracellular accommodation of arbuscules and reveal that VPY functions in both the cytoplasm and nucleus. VPY interacts with two kinases, including DOES NOT MAKE INFECTIONS3 (DMI3), a nuclear-localized symbiosis signaling kinase. Overexpression of VPY in a symbiosis-attenuated genetic background results in a dmi3 -like phenotype suggesting that VPY negatively influences DMI3 function. Overall, the data indicate a requirement for VPY in the nucleus and cytoplasm where it may coordinate signaling and cellular accommodation processes.
Topics: Ankyrin Repeat; Medicago truncatula; Mycorrhizae; Plant Proteins; Plant Roots; Symbiosis
PubMed: 36064777
DOI: 10.1038/s41467-022-32124-3 -
Cell Metabolism Oct 2019The discovery of prolyl hydroxylase domain proteins (PHDs) as key enzymes in the hypoxia inducible factor (HIF) pathway has been followed by reports of a multitude of...
The discovery of prolyl hydroxylase domain proteins (PHDs) as key enzymes in the hypoxia inducible factor (HIF) pathway has been followed by reports of a multitude of non-HIF substrates of PHD. Reporting in eLife, Cockman et al. (2019) find a surprising lack of detectable PHD activity toward any of them.
Topics: Ankyrin Repeat; Humans; Hydroxylation; Hypoxia; Procollagen-Proline Dioxygenase; Proteomics
PubMed: 31577931
DOI: 10.1016/j.cmet.2019.08.008 -
Biomolecules Jun 2021Ankyrin repeat proteins are found in all three kingdoms of life. Fundamentally, these proteins are involved in protein-protein interaction in order to activate or... (Review)
Review
Ankyrin repeat proteins are found in all three kingdoms of life. Fundamentally, these proteins are involved in protein-protein interaction in order to activate or suppress biological processes. The basic architecture of these proteins comprises repeating modules forming elongated structures. Due to the lack of long-range interactions, a graded stability among the repeats is the generic properties of this protein family determining both protein folding and biological function. Protein folding intermediates were frequently found to be key for the biological functions of repeat proteins. In this review, we discuss most recent findings addressing this close relation for ankyrin repeat proteins including DARPins, Notch receptor ankyrin repeat domain, IκBα inhibitor of NFκB, and CDK inhibitor p19. The role of local folding and unfolding and gradual stability of individual repeats will be discussed during protein folding, protein-protein interactions, and post-translational modifications. The conformational changes of these repeats function as molecular switches for biological regulation, a versatile property for modern drug discovery.
Topics: Animals; Ankyrin Repeat; Biological Phenomena; Humans; Protein Folding; Protein Stability; Protein Structure, Secondary
PubMed: 34198779
DOI: 10.3390/biom11060840