Synthetic biology principles for the design of protein with novel structures and functions

Abstract

Nature provides a large number of functional proteins that evolved during billions of years of evolution. The diversity of natural proteins encompasses versatile functions and more than a thousand different folds, which, however, represents only a tiny fraction of all possible folds and polypeptide sequences. Recent advances in the rational design of proteins demonstrate that it is possible to design de novo protein folds unseen in nature. Novel protein topologies have been designed based on similar principles as natural proteins using advanced computational modelling or modular construction principles, such as oligomerization domains. Designed proteins exhibit several interesting features such as extreme stability, designability of 3D topologies and folding pathways. Moreover, designed protein assemblies can implement symmetry similar to the viral capsids, while, on the other hand, single-chain pseudosymmetric designs can address each position independently. Recently, the design is expanding towards the introduction of new functions into designed proteins, and we may soon be able to design molecular machines.

Functionalization of a symmetric protein scaffold: Redundant folding nuclei and alternative oligomeric folding pathways

“…Protein design offers an infinite possibility of novel protein structures and functionalities with diverse application in the areas of novel therapeutics, diagnostics, sensors, materials, etc. One common approach is computational protein design targeting enzyme engineering, protein specificity, cellular pathway control, and higher‐order protein assembly 1–6 . However, as a single step, computational design has the challenging requirement of solving at least three design problems simultaneously, including (a) protein foldability (i.e., folding kinetics requirements), (b) protein stability (i.e., thermodynamic requirements), and (c) the accommodation of specific function (with potential structural dynamics requirements).…”

“…One common approach is computational protein design targeting enzyme engineering, protein specificity, cellular pathway control, and higher-order protein assembly. [1][2][3][4][5][6] However, as a single step, computational design has the challenging requirement of solving at least three design problems simultaneously, including (a) protein foldability (i.e., folding kinetics requirements), (b) protein stability (i.e., thermodynamic requirements), and (c) the accommodation of specific function (with potential structural dynamics requirements). These discrete design requirements are interconnected and likely orthogonal, as there is evidence for both a "function/stability tradeoff," 7,8 and "function/foldability tradeoff."…”

Development of mammalian cell logic gates controlled by unnatural amino acids

“…Employing logic gates in mammalian cells is a fast-developing area for intricate reversible cellular control, with significant biotechnological and biomedical applications ( Cuthbertson and Nodwell, 2013 ; Fink et al., 2019 ; Nguyen et al., 2016 ; Ross et al., 2016 ; Wu et al., 2015 ; Zetsche et al., 2015 ). Some examples include novel sensors, diagnostics, as well as therapeutics ( Brown et al., 2018 ; Kitada et al., 2018 ; Scheller and Fussenegger, 2019 ; Singh, 2014 ; Zhou et al., 2020 ). In particular, logic gates responding to small molecules can provide (spatio)temporal control by the user, permitting targeted manipulation of a designated phenotype.…”

Triangular in Vivo Self-Assembling Coiled-Coil Protein Origami

“…However, compared to DNA, polypeptides as the nanostructure building material offer greater structural and functional diversity, because they are composed of 20 amino acids with different chemical properties. In addition, proteins can be produced cost-effectively on a large scale using biotechnological methods, which make proteins attractive for designed bionanomaterials . Several strategies have been developed for the design of new protein structures based on techniques such as directed evolution, , introduction of interaction surfaces on natural oligomerization domains, − fusion of oligomerization domains with matching symmetries, − and assembly of secondary structure modules. − A coiled-coil (CC) dimer is an appealing building module for de novo polypeptide nanostructures.…”