title: Molecular dynamics simulations elucidate the mode of protein recognition by Skp1 and the F-box domain in the SCF complex.
abstract: Polyubiquitination of the target protein by a ubiquitin transferring machinery is key to various cellular processes. E3 ligase Skp1-Cul1-F-box (SCF) is one such complex which plays crucial role in substrate recognition and transfer of the ubiquitin molecule. Previous computational studies have focused on S-phase kinase-associated protein 2 (Skp2), cullin, and RING-finger proteins of this complex, but the roles of the adapter protein Skp1 and F-box domain of Skp2 have not been determined. Using sub-microsecond molecular dynamics simulations of full-length Skp1, unbound Skp2, Skp2-Cks1 (Cks1: Cyclin-dependent kinases regulatory subunit 1), Skp1-Skp2, and Skp1-Skp2-Cks1 complexes, we have elucidated the function of Skp1 and the F-box domain of Skp2. We found that the L16 loop of Skp1, which was deleted in previous X-ray crystallography studies, can offer additional stability to the ternary complex via its interactions with the C-terminal tail of Skp2. Moreover, Skp1 helices H6, H7, and H8 display vivid conformational flexibility when not bound to Skp2, suggesting that these helices can recognize and lock the F-box proteins. Furthermore, we observed that the F-box domain could rotate (5°-129°), and that the binding partner determined the degree of conformational flexibility. Finally, Skp1 and Skp2 were found to execute a domain motion in Skp1-Skp2 and Skp1-Skp2-Cks1 complexes that could decrease the distance between ubiquitination site of the substrate and the ubiquitin molecule by 3 nm. Thus, we propose that both the F-box domain of Skp2 and Skp1-Skp2 domain motions displaying preferential conformational control can together facilitate polyubiquitination of a wide variety of substrates.
authors.affiliation: Cactus Communications Pvt. Ltd, Andheri (W), Mumbai, 400053, India.|Department of Biosciences and Bioengineering, Indian Institute of Technology Bombay, Powai, Mumbai, 400076, India.
authors.forename: Sarath
authors.initials: S
authors.lastname: Chandra Dantu
authors.affiliation: Department of Biosciences and Bioengineering, Indian Institute of Technology Bombay, Powai, Mumbai, 400076, India.
authors.forename: Nitin
authors.initials: N
authors.lastname: Nathubhai Kachariya
authors.affiliation: Department of Biosciences and Bioengineering, Indian Institute of Technology Bombay, Powai, Mumbai, 400076, India.
authors.forename: Ashutosh
authors.initials: A
authors.lastname: Kumar
mesh_terms: D044842:Cullin Proteins / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D056004:Molecular Dynamics Simulation; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D044786:S-Phase Kinase-Associated Proteins / Q000737:chemistry* / Q000378:metabolism; D044767:Ubiquitin-Protein Ligases / Q000737:chemistry* / Q000378:metabolism; D054875:Ubiquitination
chemical_list: C100318:Cullin 1; D044842:Cullin Proteins; D044786:S-Phase Kinase-Associated Proteins; C097351:SKP1 protein, human; D044767:Ubiquitin-Protein Ligases
keywords: F-box rotation; conformational sampling; flexible helices; ubiquitination