Universe of Lasso Proteins: Exploring the limit of entanglement and folding landscape of proteins predicted by AlphaFold

AO_SCPLOWBSTRACTC_SCPLOWKnots and lasso topology represent a class of natural motifs found in proteins which are characterized by a threaded structure. Proteins with a lasso motif represent a macroscopic version of the peptide lasso, which are known for their high stability and offer tremendous potential for the development of novel therapeutics. Here, based on AlphaFold, we have shown the limit of topological complexity of naturally occurring protein structures with cysteine bridges. Based on 176 million high confidence (pLDDT > 70) AlphaFold-predicted protein models and a detailed analysis of the conservation of the motif in a family, we found four new lasso motifs, including L4 and LS4LS3 topologies, and the first examples of knotted lasso proteins: L1K31 and L3#K31. We show that in the case of natural proteins, there are no lassos with 5 threadings but there exist some with 6. Families possessing proteins with more than 6 threadings did not exceed the conservation threshold of 10%. Moreover, we propose a probable folding mechanism for the LS4LS3 lasso motif, enhancing our view on protein folding and stability. This work expands the topological space of lasso type motifs in proteins but also suggests that more complex structures could be unfavorable for proteins. HighlightsO_LIDiscovery of novel non-trivial lasso motifs: the L4, supercoiling of both tails LS4LS3, and the first knotted lasso proteins: L1K31 and L3#K31. C_LIO_LIThe knotted lasso motifs are in membrane proteins. C_LIO_LILassos topologies with 5 or more crossings are not conserved in protein families, and more complex motifs do not exist C_LIO_LI472 new InterPro entries with a high probability of non-trivial lasso motif C_LIO_LIPotential folding pathway for proteins with complex supercoiled lasso motif LS4LS3 C_LI Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=71 SRC="FIGDIR/small/644650v1_ufig1.gif" ALT="Figure 1"> View larger version (20K): org.highwire.dtl.DTLVardef@9c3cedorg.highwire.dtl.DTLVardef@163f941org.highwire.dtl.DTLVardef@811e31org.highwire.dtl.DTLVardef@e53de5_HPS_FORMAT_FIGEXP M_FIG C_FIG