Designing repeat proteins: a modular approach to protein design

Abstract

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.

Comparing proteins and nucleic acids for next-generation biomolecular engineering

“…DARPins have been used as crystallization chaperone 82 and sensors of protein conformation, and to induce apoptosis in tumors 83 . The concept of repeatbased recognition scaffolds is common and has been implemented using sequences with leucine-rich-repeats 84 , a 42-residue tetratricopeptide repeat variant 85 , HEAT-like repeats 86 , and others 87,88 . Figure 2C) even though the chemical diversity of endogenous nucleic acids is lower than for proteins [21][22][23][24]45 .…”

Hierarchical design of artificial proteins and complexes toward synthetic structural biology

“…Repeat proteins are defined by the repetition of a varying number of small structural units (20-50 amino acids). The modular and highly repetitive structures suggest that new artificial repeat protein repertoires can be constructed by concatenation of repetitive modules as idealized building blocks (Parmeggiani and Huang 2017). Various libraries of artificially designed repeat proteins have been generated to select…”

“…An increasing number of alternative scaffolds for artificial binding proteins have been developed using protein engineering and combinatorial approaches: monobodies derived from fibronectin type III, anticalins derived from lipocalins, affibodies derived from the immunoglobulin binding protein A, and DARPins based on the ankyrin fold can be regarded as the established formats of alternative scaffolds (Gilbreth and Koide 2012; Jost and Pluckthun 2014). Especially, the modular and highly repetitive structures of repeat proteins suggest that new artificial repeat protein repertoires can be generated by concatenation of idealized structural modules as building blocks (Boersma and Pluckthun 2011;Urvoas et al 2012;Parmeggiani and Huang 2017). Repeated structural motifs stack together to generate large protein surfaces usually involved in protein-protein interactions.…”

Elfin: An algorithm for the computational design of custom three-dimensional structures from modular repeat protein building blocks

“…In these cases, complexity is not simply reduced by constraining structurally equivalent positions to the same amino acid. Contacts between residues along the chain are limited, since generally each repeat interacts only locally with its two neighboring units, located before and after it in the primary sequence (Javadi and Itzhaki, 2013;Kajava, 2012;Paladin et al, 2017;Parmeggiani and Huang, 2017). Combinations of different repeats with compatible interfaces allow control of the overall protein shape (Park et al, 2015).…”