Computational design of a self-assembling symmetrical β-propeller protein

Abstract

The modular structure of many protein families, such as β-propeller proteins, strongly implies that duplication played an important role in their evolution, leading to highly symmetrical intermediate forms.Previous attempts to create perfectly symmetrical propeller proteins have failed, however. We have therefore developed a new and rapid computational approach to design such proteins. As a test case, we have created a sixfold symmetrical β-propeller protein and experimentally validated the structure using X-ray crystallography. Each blade consists of 42 residues. Proteins carrying 2-10 identical blades were also expressed and purified. Two or three tandem blades assemble to recreate the highly stable sixfold symmetrical architecture, consistent with the duplication and fusion theory. The other proteins produce different monodisperse complexes, up to 42 blades (180 kDa) in size, which self-assemble according to simple symmetry rules. Our procedure is suitable for creating nano-building blocks from different protein templates of desired symmetry.protein evolution | computational protein design | self-assembly | β-propeller | protein crystallography

CODV-Ig, a universal bispecific tetravalent and multifunctional immunoglobulin format for medical applications

“…The CODV design is thus akin to the design of circularly closed repeat architectures such as b-propeller and b-trefoil folds. 56,[63][64][65] Steric constraints must be matched to permit correct VH/VL pairing of Fv1 and Fv2, as in engineering of circularly closed repeat architectures. Inappropriate linker lengths may result in VH/VL mismatch, leading to dimerization of DVD-Ig type with loss of target affinity as suggested by our initial constructs (Supplements).…”

Designed protein reveals structural determinants of extreme kinetic stability

“…These features may be "inherited" through the use of existing sequence information, generating a more naturally funneled energy landscape. Other proteins designed in a similar way, and not yet characterized in detail, may also capture favorable natural features (16)(17)(18). Also, our results show how ligand binding can further smooth the landscape by decreasing the formation of nonnative structure and so promote folding and design success.…”

“…Design strategies similar to that used to make ThreeFoil, which use repetition of structural elements designed using existing sequence information and structural modeling with the Rosetta energy function (28), have proven particularly fruitful, with several studies yielding well-folded proteins with high melting points on the first attempt (1,(16)(17)(18). Furthermore, we have shown that ThreeFoil possesses stability, cooperativity and multivalent binding function.…”

“…Internal structural symmetry, resulting from the repetition of smaller elements of structure, is very common in natural proteins, with ∼20% of all protein folds (22) and the majority of the most populated globular protein folds (superfolds) (21) containing internal structural symmetry. Recent design successes, for helical proteins (5, 6), repeat proteins (18,20,23) and symmetric superfolds (1,2,7,16,17,19,(24)(25)(26) recommend the simplification of the design process by using repetitive/symmetric folds as a particularly effective strategy.…”

Internal symmetry in protein structures: prevalence, functional relevance and evolution

“…Attempts to elucidate evolutionary models on a selected set of symmetric protein domains such as b-trefoils and b-propellers have been successful [7 ,24]. This inspired the design of symmetric protein structures with a potential for various applications [25][26][27]. Particularly in the last five years, there has been significant progress in identifying protein structural symmetry and describing its relevance to functions and evolution of proteins, which will be the primary focus below.…”