Location dependent coordination chemistry and MRI relaxivity, in de novo designed lanthanide coiled coils

AbstractMango is an economically important fruit crop largely cultivated in the (sub)tropics and thus, is constantly challenged by a myriad of insect pests and diseases. Here, we identified and characterized the resistance gene analogs (RGAs) of mango from de novo assembly of transcriptomic sequences. A core RGA database of mango with 747 protein models was established and classified based on conserved domains and motifs: 53 nucleotide binding site proteins (NBS); 27 nucleotide binding site-leucine rich repeat proteins (NBS-LRR); 17 coiled-coil NBS-LRR (CNL); 2 toll/interleukin-1 receptor NBS-LRR (TNL); 29 coiled-coil NBS (CN); 4 toll/interleukin-1 receptor NBS (TN); 17 toll/interleukin-1 receptor with unknown domain (TX); 158 receptor-like proteins (RLP); 362 receptor-like kinases (RLK); 72 transmembrane coiled-coil domain protein (TM-CC), and 6 NBS-encoding proteins with other domains. The various molecular functions, biological processes, and cellular localizations of these RGAs were functionally well-annotated through gene ontology (GO) analysis, and their expression profiles across different mango varieties were also determined. Phylogenetic analysis broadly clustered the core RGAs into 6 major clades based on their domain classification, while TM-CC proteins formed subclades all across the tree. The phylogenetic results suggest highly divergent functions of the RGAs which also provide insights into the mango-pest co-evolutionary arms race. From the mango RGA transcripts, 134 unique EST-SSR loci were identified, and primers were designed targeting these potential markers. To date, this is the most comprehensive analysis of mango RGAs which offer a trove of markers for utilization in resistance breeding of mango.

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De novo design of α-helical proteins: basic research to medical applications

Biochemistry and Cell Biology ◽

10.1139/o96-015 ◽

1996 ◽

Vol 74 (2) ◽

pp. 133-154 ◽

Cited By ~ 106

Author(s):

Robert S. Hodges

Keyword(s):

Protein Folding ◽

De Novo ◽

Coiled Coil ◽

Basic Research ◽

De Novo Design ◽

Coiled Coils ◽

Medical Applications ◽

Dimerization Domain ◽

Recombinant Peptides ◽

Helical Proteins

The two-stranded α-helical coiled-coil is a universal dimerization domain used by nature in a diverse group of proteins. The simplicity of the coiled-coil structure makes it an ideal model system to use in understanding the fundamentals of protein folding and stability and in testing the principles of de novo design. The issues that must be addressed in the de novo design of coiled-coils for use in research and medical applications are (i) controlling parallel versus antiparallel orientation of the polypeptide chains, (ii) controlling the number of helical strands in the assembly (iii) maximizing stability of homodimers or heterodimers in the shortest possible chain length that may require the engineering of covalent constraints, and (iv) the ability to have selective heterodimerization without homodimerization, which requires a balancing of selectivity versus affinity of the dimerization strands. Examples of our initial inroads in using this de novo design motif in various applications include: heterodimer technology for the detection and purification of recombinant peptides and proteins; a universal dimerization domain for biosensors; a two-stage targeting and delivery system; and coiled-coils as templates for combinatorial helical libraries for basic research and drug discovery and as synthetic carrier molecules. The universality of this dimerization motif in nature suggests an endless number of possibilities for its use in de novo design, limited only by the creativity of peptide–protein engineers.Key words: de novo design of proteins, α-helical coiled-coils, protein folding, protein stability, dimerization domain, dimerization motif.

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De novo design of isopeptide bond-tethered triple-stranded coiled coils with exceptional resistance to unfolding and proteolysis: implication for developing antiviral therapeutics

Chemical Science ◽

10.1039/c5sc02220g ◽

2015 ◽

Vol 6 (11) ◽

pp. 6505-6509 ◽

Cited By ~ 6

Author(s):

Chao Wang ◽

Wenqing Lai ◽

Fei Yu ◽

Tianhong Zhang ◽

Lu Lu ◽

...

Keyword(s):

De Novo ◽

Coiled Coil ◽

De Novo Design ◽

Coiled Coils ◽

Isopeptide Bond ◽

Hiv 1

Isopeptide bridge-tethered ultra-stable coiled-coil trimers have been de novo designed as structure-directing auxiliaries to guide HIV-1 gp41 NHR-peptide trimerization.

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De novo design of a non-natural fold for an iron–sulfur protein: Alpha-helical coiled-coil with a four-iron four-sulfur cluster binding site in its central core

Biochimica et Biophysica Acta (BBA) - Bioenergetics ◽

10.1016/j.bbabio.2009.12.012 ◽

2010 ◽

Vol 1797 (3) ◽

pp. 406-413 ◽

Cited By ~ 46

Author(s):

Joanna Grzyb ◽

Fei Xu ◽

Lev Weiner ◽

Eduard J. Reijerse ◽

Wolfgang Lubitz ◽

...

Keyword(s):

Binding Site ◽

De Novo ◽

Coiled Coil ◽

De Novo Design ◽

Central Core ◽

Sulfur Cluster ◽

Iron Sulfur ◽

Sulfur Protein ◽

Iron Sulfur Protein

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Coiled coils 9-to-5: Rational de novo design of α-helical barrels with tunable oligomeric states

10.1101/2021.01.20.427391 ◽

2021 ◽

Author(s):

William M. Dawson ◽

Freddie J.O. Martin ◽

Guto G. Rhys ◽

Kathryn L. Shelley ◽

R. Leo Brady ◽

...

Keyword(s):

Crystal Structures ◽

Rational Design ◽

De Novo ◽

Coiled Coil ◽

De Novo Design ◽

Coiled Coils ◽

Alternative States ◽

X Ray ◽

Amphipathic Helices ◽

Made In

ABSTRACTThe rational design of linear peptides that assemble controllably and predictably in water is challenging. Sequences must encode unique target structures and avoid alternative states. However, the stabilizing and discriminating non-covalent forces available are weak in water. Nonetheless, for α-helical coiled-coil assemblies considerable progress has been made in rational de novo design. In these, sequence repeats of nominally hydrophobic (h) and polar (p) residues, hpphppp, direct the assembly of amphipathic helices into dimeric to tetrameric bundles. Expanding this pattern to hpphhph can produce larger α-helical barrels. Here, we show that pentamers to nonamers are achieved simply by varying the residue at one of these h sites. In L/I-K-E-I-A-x-Z repeats, decreasing the size of Z from threonine to serine to alanine to glycine gives progressively larger oligomers. X-ray crystal structures of the resulting α-helical barrels rationalize this: side chains at Z point directly into the helical interfaces, and smaller residues allow closer helix contacts and larger assemblies.

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Automatedde novophasing and model building of coiled-coil proteins

Acta Crystallographica Section D Biological Crystallography ◽

10.1107/s1399004714028247 ◽

2015 ◽

Vol 71 (3) ◽

pp. 606-614 ◽

Cited By ~ 12

Author(s):

Sebastian Rämisch ◽

Robert Lizatović ◽

Ingemar André

Keyword(s):

Structure Prediction ◽

Initial Phase ◽

Model Building ◽

De Novo ◽

Protein Complexes ◽

Coiled Coil ◽

Coiled Coils ◽

Phase Estimation ◽

Molecular Replacement ◽

Experimental Phasing

Models generated byde novostructure prediction can be very useful starting points for molecular replacement for systems where suitable structural homologues cannot be readily identified. Protein–protein complexes andde novo-designed proteins are examples of systems that can be challenging to phase. In this study, the potential ofde novomodels of protein complexes for use as starting points for molecular replacement is investigated. The approach is demonstrated using homomeric coiled-coil proteins, which are excellent model systems for oligomeric systems. Despite the stereotypical fold of coiled coils, initial phase estimation can be difficult and many structures have to be solved with experimental phasing. A method was developed for automatic structure determination of homomeric coiled coils from X-ray diffraction data. In a benchmark set of 24 coiled coils, ranging from dimers to pentamers with resolutions down to 2.5 Å, 22 systems were automatically solved, 11 of which had previously been solved by experimental phasing. The generated models contained 71–103% of the residues present in the deposited structures, had the correct sequence and had freeRvalues that deviated on average by 0.01 from those of the respective reference structures. The electron-density maps were of sufficient quality that only minor manual editing was necessary to produce final structures. The method, namedCCsolve, combines methods forde novostructure prediction, initial phase estimation and automated model building into one pipeline.CCsolveis robust against errors in the initial models and can readily be modified to make use of alternative crystallographic software. The results demonstrate the feasibility ofde novophasing of protein–protein complexes, an approach that could also be employed for other small systems beyond coiled coils.

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Navigating the structural landscape of de novo α–helical bundles

10.1101/503698 ◽

2018 ◽

Author(s):

Guto G. Rhys ◽

Christopher W. Wood ◽

Joseph L. Beesley ◽

Nathan R. Zaccai ◽

Antony J. Burton ◽

...

Keyword(s):

De Novo ◽

Hydrophobic Effect ◽

Protein Structures ◽

Coiled Coil ◽

Coiled Coils ◽

Single Amino Acid ◽

Helical Bundles ◽

Helical Bundle ◽

Peptide System ◽

Antiparallel Coiled Coil

ABSTRACTThe association of amphipathic α helices in water leads to α-helical-bundle protein structures. However, the driving force for this—the hydrophobic effect—is not specific and does not define the number or the orientation of helices in the associated state. Rather, this is achieved through deeper sequence-to-structure relationships, which are increasingly being discerned. For example, for one structurally extreme but nevertheless ubiquitous class of bundle—the α-helical coiled coils—relationships have been established that discriminate between all-parallel dimers, trimers and tetramers. Association states above this are known, as are antiparallel and mixed arrangements of the helices. However, these alternative states are less-well understood. Here, we describe a synthetic-peptide system that switches between parallel hexamers and various up-down-up-down tetramers in response to single-amino-acid changes and solution conditions. The main accessible states of each peptide variant are characterized fully in solution and, in most cases, to high-resolution X-ray crystal structures. Analysis and inspection of these structures helps rationalize the different states formed. This navigation of the structural landscape of α-helical coiled coils above the dimers and trimers that dominate in nature has allowed us to design rationally a well-defined and hyperstable antiparallel coiled-coil tetramer (apCC-Tet). This robust de novo protein provides another scaffold for further structural and functional designs in protein engineering and synthetic biology.

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Design of two-stranded and three-stranded coiled-coil peptides

Philosophical Transactions of the Royal Society B Biological Sciences ◽

10.1098/rstb.1995.0048 ◽

1995 ◽

Vol 348 (1323) ◽

pp. 81-88 ◽

Cited By ~ 52

Keyword(s):

Protein Design ◽

De Novo ◽

Coiled Coil ◽

Structural Features ◽

Coiled Coils ◽

General Sequence ◽

The Third ◽

Cooperative Equilibrium ◽

High Concentrations ◽

Propensity Scale

The structural features required for the formation of two- versus three-stranded coiled coils have been explored using de novo protein design. Peptides with leucine at the ‘a’ and ‘d’ positions of a coiled-coil (general sequence: Leu a Xaa b Xaa c Leu d Glu e Xaa f Lys g ) exist in a non-cooperative equilibrium between unstructured monomers and helical dimers and helical trimers. Substituting valine into each ‘a’ position produces peptides which still form trimers at high concentrations, whereas substitution of a single asparagine at the ‘a’ position of the third heptad yields a dimer. During the course of this work, we also re-investigated a helical propensity scale derived using a series of coiled-coil peptides previously believed to exist in a monomer-dimer equilibrium (O’Neil & DeGrado 1990). Detailed analysis of the concentration dependence of ellipticity at 222 nm reveals that they exist in a non-cooperative monomer-dimer-trimer equilibrium. However, the concentration of trimer near the midpoint of the concentration-dependent transition is small, so the previously determined values of ΔΔG α using the approximate monomer-dimer scheme are indistinguishable from the values obtained employing the complete monomer-dimer-trimer equilibrium.

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Metal Ion Assisted Folding and Supramolecular Organization of a De Novo Designed Metalloprotein

Australian Journal of Chemistry ◽

10.1071/ch03260 ◽

2004 ◽

Vol 57 (1) ◽

pp. 33 ◽

Cited By ~ 9

Author(s):

Guido W. M. Vandermeulen ◽

Christos Tziatzios ◽

Dieter Schubert ◽

Philip R. Andres ◽

Alexander Alexeev ◽

...

Keyword(s):

Protein Folding ◽

Self Assembly ◽

Metal Ion ◽

De Novo ◽

Coiled Coil ◽

Coiled Coils ◽

The Self ◽

Supramolecular Organization ◽

Low Concentrations ◽

The Moment

This paper describes the supramolecular organization of a novel de novo designed metalloprotein, which consists of two N-terminal terpyridine modified coiled-coil protein folding motif sequences held together by an iron(II) ion. The self-assembly of the metalloprotein is the result of the interplay of metal ion complexation and protein folding, and can be manipulated by changes in concentration, temperature, and solvent. At low concentrations, folding and organization of the metalloprotein resembles that of the native coiled-coil peptide. Besides unimeric species, also dimeric and tetrameric metalloprotein assemblies were found. Several indications suggest that at least part of these unimeric species may exist as intramolecularly folded coiled-coils, however, unambiguous proof is lacking at the moment. At higher concentrations, folding and organization is dominated by the large octahedral [FeII(terpy)2] complexes (terpy = 2,2′:6′,2″-terpyridine) and considerable amounts of large, ill-defined aggregates are formed.

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