Glycosylation Weakens Skp1 Homodimerization in Toxoplasma gondii by Interrupting a Fuzzy Interaction

Le résumé fourni par la source

High Resolution Image Download MS PowerPoint Slide Skp1/Cullin-1/F-Box protein (SCF) complexes represent a major class of E3 ubiquitin ligases responsible for proteomic control throughout eukaryotes. Target specificity is mediated by a large set of F-box proteins (FBPs) whose F-box domains interact with Skp1 in a conserved, well-organized fashion. In the social amoeba Dictyostelium, Skp1 is regulated by oxygen-dependent glycosylation which alters Skp1’s FBP interactome and inhibits homodimerization that is mediated in part by an ordered interface which overlaps with that of FBPs. Based on sedimentation velocity experiments, Skp1 from the intracellular pathogen Toxoplasma gondii exhibits a homodimerization K d comparable to that of a previously measured FBP/Skp1 interaction. Glycosylation of Skp1’s disordered C-terminal region (CTR) distal to the ordered homodimer interface significantly weakens Skp1 homodimerization, an effect reproduced by CTR deletion. Replacement with a randomized CTR sequence retains high affinity excluding an extension of the ordered dimer interface. Substitution by poly serine weakens the homodimer to a degree equal to its deletion, indicating a composition dependent effect. The contribution of the CTR to Skp1 homodimerization is canceled by high salt consistent with an electrostatic mechanism. All-atom molecular dynamics simulations suggest that the CTR promotes homodimerization via charge cluster interactions. Taken together, the data indicate that glycosylation weakens homodimerization by disrupting a C-terminal fuzzy interaction that functions in tandem with the ordered dimer interface, thereby freeing Skp1 for FBP binding. Thus, the CTR contributes to Skp1/Skp1 and Skp1/FBP interactions via independent mechanisms that are each influenced by O 2, indicating multiple constraints on the evolution of its sequence.

Ce résumé expose les affirmations des auteurs. BNTIC ne l’interprète pas comme une validation indépendante des résultats.