Designed Ankyrin Repeat Proteins as Scaffolds for Multivalent Recognition

Abstract

Ankyrin repeat (AR) proteins are composed of tandem repeats of a basic structural motif of ca. 33 amino acid residues that form a β-turn followed by two antiparallel α-helices. Multiple repeats stack together in a modular fashion to form a scaffold that is ideally suited for the presentation of multiple functional groups and/or recognition elements. Here we describe a biosynthetic strategy that takes advantage of the modular nature of these proteins to generate multivalent ligands that are both chemically homogeneous and structurally well-defined. Glycosylated AR proteins cluster the tetrameric lectin concanavalin A (Con A) at a rate that is comparable to the rate of Con A aggregation mediated by globular protein conjugates and variable density linear polymers. Thus, AR proteins define a new class of multivalent ligand scaffolds that have significant potential application in the study and control of a variety of multivalent interactions.

Conserved Ankyrin Repeat Proteins and Their NIMA Kinase Partners Regulate Extracellular Matrix Remodeling and Intracellular Trafficking inCaenorhabditis elegans

“…Splicing from exon 5 to exon 7 in sv9 mutants would be predicted to eliminate the fifth ankyrin repeat, as well as parts of the fourth and sixth repeats, but would not induce a frameshift. Ankyrin repeat domains are important for protein-protein interactions and can serve as molecular scaffolds (Mosavi et al 2004;Voronin and Kiseleva 2008;Hollenbeck et al 2012). Based on reciprocal BLAST and OrthoList (Shaye and Greenwald 2011), MLT-4 is orthologous to human inversin (INVS), and more similar in amino acid sequence than a previously reported INVS ortholog, NPHP-2 ( Figure S1 and Figure S2) (Warburton-Pitt et al 2012).…”

A Protein‐Based Pentavalent Inhibitor of the Cholera Toxin B‐Subunit

“…As the majority of the structure comes as a ready-made building block, the synthetic route to the inhibitor is relatively short. Previous attempts to introduce sugars at defined sites on an ankyrin repeat protein[ 10 ] and the barstar protein[ 11 ] led to ligands and inhibitors of plant lectins. However, as no attempts were made to match the size and spacing of the ligands to the binding sites, these multivalent compounds led to only modest enhancements in activity and/or lectin aggregation, which may be undesirable.…”

Emerging functions for ANKHD1 in cancer-related signaling pathways and cellular processes

“…ANKHD1/p21 axis leads to progression in the cell cycle and increased pro-liferation. ( 6 ) ANKHD1 interacts with SMYD3 and assists in the tri-methylation at the 4th lysine residue of histone H3 (H3K4me3), this epigenetic alteration provides the expression of genes involved in cellular proliferation, migration, and invasion. Abbreviations: P, phosphorylation; Me, Methylation; TF, transcription factor.…”

“…Unlike other protein domains, such as the SH2 and SH3 domains, ARD does not recognize a specific amino acid sequence or protein structure, and it can interact with several other domains ( 4 , 5 ). ARD-containing proteins are found in all species and are fundamental to biological processes, including cell cycle control, transcrip-tional regulation, differentiation, apoptosis, and cytoskeletal organization ( 3 , 6 ). The KH domains bind to RNA or ssDNA, and are found in proteins associated with transcriptional and translational regulation ( 7 - 9 ).…”