AlphaFold predicts novel human proteins with knots

Abstract

The fact that proteins can have their chain formed in a knot is known for almost 30 years. However, as they are not common, only a fraction of such proteins is available in the Protein Data Bank. It was not possible to assess their importance and versatility up until now because we did not have access to the whole proteome of an organism, let alone a human one. The arrival of efficient machine learning methods for protein structure prediction, such as AlphaFold and RoseTTaFold, changed that. We analyzed all proteins from the human proteome (over 20,000) determined with AlphaFold in search for knots and found them in less than 2% of the structures. Using a variety of methods, including homolog search, clustering, quality assessment, and visual inspection, we determined the nature of each of the knotted structures and classified it as either knotted, potentially knotted, or an artifact, and deposited all of them in a database available at: https://knotprot.cent.uw.edu.pl/alphafold. Overall, we found 51 credible knotted proteins (0.2% of human proteome). The set of potentially knotted structures includes a new complex type of a knot not reported in proteins yet. That knot type, denoted 6₃ in mathematical notation, would necessitate a more complex folding path than any knotted protein characterized to date.

Keywords: biological function; evolution; folding; new knotted folds.

FIGURE 1

Selected proteins with a 3₁ knot. (a) Simplified representation of a 3₁ knot found in proteins. (b) Ecdysoneless protein (UniProtKB: O95905). (c) Calcium‐activated chloride channel regulator 1 (UniProtKB ID: A8K7I4). (d) Integrin alpha IIb (UniProtKB ID: P08514). All knotted cores are shown in a rainbow color scheme to guide the eye. (e) Upper panel: unknotted integrin alpha 1 (UniProtKB ID: P56199). Lower panel: 3₁ knotted integrin alpha 3 (UniProtKB ID: P26006).

FIGURE 2

Von Willebrand factor A domain‐containing protein 5A (UniProtKB ID: O00534) with a potential new type of protein knot—6₃ shown with essential strands colored in red, blue, and purple. Upper left panel shows a simplified knotted core of the protein.