Modular Evolution and the Origins of Symmetry: Reconstruction of a Three-Fold Symmetric Globular Protein

Broom, Aron; Doxey, Andrew C.; Lobsanov, Y.D.; Berthin, Lisa G.; Rose, David R.; Howell, P. Lynne; McConkey, Brendan J.; Meiering, Elizabeth M.

doi:10.1016/j.str.2011.10.021

Cited by 99 publications

(103 citation statements)

References 64 publications

(102 reference statements)

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“…56 We note that a protein with folding pathway redundancy might, in principle, retain efficient folding despite a localized deleterious mutation due to compensation by the folding-competent symmetry-related positions. Recent reports of de novo designed purely symmetric b-trefoil proteins that efficiently fold and are hyperthermophile in stability [57][58][59] demonstrate that pure primary structure symmetry does not necessarily result in folding frustration and therefore supports the hypothesis of redundant foldability. If folding redundancy is a property of purely symmetric proteins (i.e., as would result from gene duplication and fusion replication errors [60][61][62] ), it highlights a critical advantage of symmetric protein architecture over asymmetric architecture in protein evolution and de novo design; namely, an intrinsic ability to tolerate diverse functional mutation and retain efficient foldability.…”

Section: Discussionmentioning

confidence: 86%

Experimental support for the foldability–function tradeoff hypothesis: Segregation of the folding nucleus and functional regions in fibroblast growth factor‐1

2012

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The acquisition of function is often associated with destabilizing mutations, giving rise to the stability-function tradeoff hypothesis. To test whether function is also accommodated at the expense of foldability, fibroblast growth factor-1 (FGF-1) was subjected to a comprehensive u-value analysis at each of the 11 turn regions. FGF-1, a b-trefoil fold, represents an excellent model system with which to evaluate the influence of function on foldability: because of its threefold symmetric structure, analysis of FGF-1 allows for direct comparisons between symmetryrelated regions of the protein that are associated with function to those that are not; thus, a structural basis for regions of foldability can potentially be identified. The resulting u-value distribution of FGF-1 is highly polarized, with the majority of positions described as either foldedlike or denatured-like in the folding transition state. Regions important for folding are shown to be asymmetrically distributed within the protein architecture; furthermore, regions associated with function (i.e., heparin-binding affinity and receptor-binding affinity) are localized to regions of the protein that fold after barrier crossing (late in the folding pathway). These results provide experimental support for the foldability-function tradeoff hypothesis in the evolution of FGF-1. Notably, the results identify the potential for folding redundancy in symmetric protein architecture with important implications for protein evolution and design.

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Section: Discussionmentioning

confidence: 86%

Experimental support for the foldability–function tradeoff hypothesis: Segregation of the folding nucleus and functional regions in fibroblast growth factor‐1

2012

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“…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).…”

Section: Discussionmentioning

confidence: 99%

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

Steinmetz

Vallée

Beil

et al. 2016

mAbs

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Bispecific immunoglobulins (Igs) typically contain at least two distinct variable domains (Fv) that bind to two different target proteins. They are conceived to facilitate clinical development of biotherapeutic agents for diseases where improved clinical outcome is obtained or expected by combination therapy compared to treatment by single agents. Almost all existing formats are linear in their concept and differ widely in drug-like and manufacture-related properties. To overcome their major limitations, we designed cross-over dual variable Ig-like proteins (CODV-Ig). Their design is akin to the design of circularly closed repeat architectures. Indeed, initial results showed that the traditional approach of utilizing (G4S) x linkers for biotherapeutics design does not identify functional CODV-Igs. Therefore, we applied an unprecedented molecular modeling strategy for linker design that consistently results in CODV-Igs with excellent biochemical and biophysical properties. CODV architecture results in a circular self-contained structure functioning as a self-supporting truss that maintains the parental antibody affinities for both antigens without positional effects. The format is universally suitable for therapeutic applications targeting both circulating and membrane-localized proteins. Due to the full functionality of the Fc domains, serum half-life extension as well as antibody-or complement-dependent cytotoxicity may support biological efficiency of CODV-Igs. We show that judicious choice in combination of epitopes and paratope orientations of bispecific biotherapeutics is anticipated to be critical for clinical outcome. Uniting the major advantages of alternative bispecific biotherapeutics, CODV-Igs are applicable in a wide range of disease areas for fast-track multi-parametric drug optimization.

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“…Templates used can range from a single repeat unit, to multiple units, to a full repeat protein domain. When a native template with highly symmetric backbone is available, designs can be carried out directly on the full structure, as shown by Broom et al [28], who generated a symmetric beta trefoil by applying a consensus sequence in the most conserved positions of a starting template and designing the rest of the sequence with the Rosetta molecular modeling suite [29].…”

Section: Structure Based Designmentioning

confidence: 99%

Designing repeat proteins: a modular approach to protein design

Parmeggiani

Huang

2017

Current Opinion in Structural Biology

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General rightsThis document is made available in accordance with publisher policies. Please cite only the published version using the reference above. Full terms of use are available: http://www.bristol.ac.uk/pure/about/ebr-terms Abstract Repeat proteins present unique opportunities for engineering because of their modular nature that potentially allows LEGO Ⓡ like construction of macromolecules. Nature takes advantage of these properties and uses this type of scaffold for recognition, structure, and even signaling purposes. In recent years, new protein modeling tools facilitated the design of novel repeat proteins, creating possibilities beyond naturally occurring scaffolds alone. We highlight here the different design strategies and summarize the various structural families and novel proteins achieved. Introduction A goal of protein engineers is to understand how sequence and structure features contribute to the makeup of proteins. However, polypeptide complexity and variability complicate the analysis of these features. Proteins that carry distinct patterns of repetitive sequences (and therefore structural features) are ideal systems with reduced complexity to inform our understanding of proteins. These repeat proteins are widespread in nature [1] and the evolutionary process that created them is quite remarkable: a segment of sequence that is structurally compatible with itself is duplicated in tandem, and the connected segments diverge to accommodate new functions within the tolerance of structural compatibility [2]. Splicing and duplication of genes govern the highly efficient and economic ways -by which higher order structures are built from recycling and repeating basic modules -to engineer structurally coupled functions. As a consequence, the repeated segments that make up a repeat protein are often not identical. However, the modularity inspires engineering efforts. In this review, we discuss the methods developed for designing repeat proteins, their achievements, and the new directions ahead.

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Modular Evolution and the Origins of Symmetry: Reconstruction of a Three-Fold Symmetric Globular Protein

Cited by 99 publications

References 64 publications

Experimental support for the foldability–function tradeoff hypothesis: Segregation of the folding nucleus and functional regions in fibroblast growth factor‐1

Experimental support for the foldability–function tradeoff hypothesis: Segregation of the folding nucleus and functional regions in fibroblast growth factor‐1

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

Designing repeat proteins: a modular approach to protein design

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