Biology subjects

Rhys, G. G.

Publications and source records attributed to Rhys, G. G..

3 recordsLinked to original sources

bioRxiv2021

How coiled-coil assemblies accommodate multiple aromatic residues

Rational protein design requires understanding the contribution of each amino acid to a targeted protein fold. For a subset of protein structures, namely the ;-helical coiled coils (CCs), knowledge is sufficiently advanced to allow the rational de novo design of many structures, including entirely new protein folds. However, current CC design rules center on using aliphatic hydrophobic residues predominantly to drive the folding and assembly of amphipathic helices. The consequences of using aromatic residues--which would be useful for introducing structural probes, and binding and catalytic functionalities--into these interfaces is not understood. There are specific examples of designed CCs containing such aromatic residues, e.g., phenylalanine-rich sequences, and the use of polar aromatic residues to make buried hydrogen-bond networks. However, it is not known generally if sequences rich in tyrosine can form CCs, or what CC assemblies these would lead to. Here we explore tyrosine-rich sequences in a general CC-forming background and resolve new CC structures. In one of these, an antiparallel tetramer, the tyrosine residues are solvent accessible and pack at the interface between the core and the surface. In the other more-complex structure, the residues are buried and form an extended hydrogen-bond network.

biochemistry↗

bioRxiv2021

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

The 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 -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 -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 -helical barrels rationalize this: side chains at Z point directly into the helical interfaces, and smaller residues allow closer helix contacts and larger assemblies.

biochemistry↗

bioRxiv2018

Navigating the structural landscape of de novo α-helical bundles

The association of amphipathic helices in water leads to -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 -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 -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.

biochemistry↗