An ankyrin repeat is a common structural motif found in proteins across nearly all forms of life, including bacteria, archaea, and eukaryotes, though they are most prevalent in eukaryotes. The structure of the repeat allows it to connect with other components to form larger, functional protein assemblies. This modular nature is the basis for the diverse roles these proteins play within the cell.
The Structure of an Ankyrin Repeat
A single ankyrin repeat is composed of approximately 33 amino acids. This sequence folds into a shape defined by two alpha-helices and a projecting beta-hairpin structure. A solitary repeat is not stable on its own; stability is achieved when multiple repeats, often four to six, are arranged in a tandem array.
These individual repeats stack together, creating an elongated structure known as an ankyrin repeat domain (ARD). This stacking is stabilized by hydrophobic interactions between the helices of adjacent repeats. The resulting architecture has an L-shaped cross-section and forms a continuous solenoid shape with two distinct surfaces: a concave face formed by the beta-hairpins and a convex face formed by the alpha-helices.
The Functional Role of Ankyrin Repeat Domains
The primary function of an ankyrin repeat domain is to mediate protein-protein interactions. Its elongated structure creates an extensive, variable surface area that allows it to bind to a wide range of different target proteins. This interaction surface does not recognize a single, universal amino acid sequence, which accounts for its versatility.
Ankyrin repeat domains act as molecular scaffolds. By binding to multiple proteins simultaneously, they bring them into close proximity to function together as part of a larger complex. This scaffolding is involved in maintaining cell structure by linking membrane proteins to the cytoskeleton, regulating the flow of information through cell signaling pathways, and controlling the progression of the cell cycle. The ankyrin repeat domain serves as a platform for assembling protein machinery, not as an enzyme with catalytic activity.
Connection to Human Disease
Since ankyrin repeat proteins are involved in many cellular activities, genetic mutations that alter their structure can impact human health. A faulty ankyrin repeat can disrupt protein binding, leading to the breakdown of cellular processes and contributing to various diseases.
A well-documented example is hereditary spherocytosis, a form of hemolytic anemia where red blood cells are misshapen, fragile, and prematurely destroyed. About half of all cases are linked to mutations in the ANK1 gene, which encodes a protein called ankyrin-1. This protein tethers the red blood cell’s membrane to its internal cytoskeleton, giving the cell its shape and durability. Mutations in ANK1’s ankyrin repeats compromise this linkage, causing cells to become spherical and rigid, leading to their destruction.
Ankyrin repeats are also involved in regulating inflammation. Proteins in the IκB family use their ankyrin repeats to bind and inhibit the transcription factor NF-κB in the cytoplasm. Upon an inflammatory stimulus, IκB is degraded, releasing NF-κB to travel to the nucleus and activate inflammatory genes. Dysregulation of this system due to mutations can cause persistent NF-κB activation, a feature of many autoimmune disorders and some cancers.
Engineered Ankyrin Repeats in Research and Therapy
Researchers have harnessed the binding properties of ankyrin repeats to create custom binding proteins known as Designed Ankyrin Repeat Proteins (DARPins). Using the natural domain as a template, these synthetic proteins are engineered in the lab to be highly stable and easy to produce in large quantities.
The creation process involves generating large libraries of DARPin variants with modified binding surfaces. From these libraries, scientists can select DARPins that bind to a specific target protein with high affinity and specificity. This makes them useful research tools for detecting the presence of a specific protein, inhibiting its function, or acting as intracellular sensors to monitor cellular events.
DARPins also represent a new class of therapeutic drugs, valued for their small size and high stability. Engineered DARPins are being developed to target disease-related proteins, such as blocking growth factor receptors on cancer cells to stop their growth. Several DARPin-based therapies are in clinical trials for treating conditions like age-related macular degeneration and various cancers.