Disorder in the Human Skp1 Structure is the Key to its Adaptability to Bind Many Different Proteins in the SCF Complex Assembly

Amrita Bhattacharya; Vaibhav Kumar Shukla; Nitin Kachariya; Preeti; Parveen Sehrawat; Ashutosh Kumar

doi:10.1016/j.jmb.2022.167830

Disorder in the Human Skp1 Structure is the Key to its Adaptability to Bind Many Different Proteins in the SCF Complex Assembly

J Mol Biol. 2022 Nov 15;434(21):167830. doi: 10.1016/j.jmb.2022.167830. Epub 2022 Sep 15.

Authors

Amrita Bhattacharya¹, Vaibhav Kumar Shukla², Nitin Kachariya¹, Preeti¹, Parveen Sehrawat¹, Ashutosh Kumar³

Affiliations

¹ Lab No. 606, Department of Biosciences and Bioengineering, Indian Institute of Technology Bombay, Powai, Mumbai 400076, India.
² Biophysical Chemistry & Structural Biology Laboratory, UM-DAE Centre for Excellence in Basic Sciences, University of Mumbai, Vidyanagari Campus, Mumbai 400098, India. Electronic address: https://twitter.com/bhu_vaibhav.
³ Lab No. 606, Department of Biosciences and Bioengineering, Indian Institute of Technology Bombay, Powai, Mumbai 400076, India. Electronic address: ashutoshk@iitb.ac.in.

PMID: 36116539
DOI: 10.1016/j.jmb.2022.167830

Abstract

Skp1(S-phase kinase-associated protein 1 - Homo sapiens) is an adapter protein of the SCF(Skp1-Cullin1-Fbox) complex, which links the constant components (Cul1-RBX) and the variable receptor (F-box proteins) in Ubiquitin E3 ligase. It is intriguing how Skp1 can recognise and bind to a variety of structurally different F-box proteins. For practical reasons, previous efforts have used truncated Skp1, and thus it has not been possible to track the crucial aspects of the substrate recognition process. In this background, we report the solution structure of the full-length Skp1 protein determined by NMR spectroscopy for the first time and investigate the sequence-dependent dynamics in the protein. The solution structure reveals that Skp1 has an architecture: β1-β2-H1-H2-L1-H3-L2-H4-H5-H6-H7(partially formed) and a long tail-like disordered C-terminus. Structural analysis using DALI (Distance Matrix Alignment) reveals conserved domain structure across species for Skp1. Backbone dynamics investigated using NMR relaxation suggest substantial variation in the motional timescales along the length of the protein. The loops and the C-terminal residues are highly flexible, and the (R₂/R₁) data suggests μs-ms timescale motions in the helices as well. Further, the dependence of amide proton chemical shift on temperature and curved profiles of their residuals indicate that the residues undergo transitions between native state and excited state. The curved profiles for several residues across the length of the protein suggest that there are native-like low-lying excited states, particularly for several C-terminal residues. Our results provide a rationale for how the protein can adapt itself, bind, and get functionally associated with other proteins in the SCF complex by utilising its flexibility and conformational sub-states.

Keywords: NMR Dynamics; SCF complex; Skp1; protein Structure; structural adaptability.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Humans
Intrinsically Disordered Proteins* / chemistry
Protein Structure, Secondary
S-Phase Kinase-Associated Proteins* / chemistry
SKP Cullin F-Box Protein Ligases* / chemistry

Substances

S-Phase Kinase-Associated Proteins
SKP Cullin F-Box Protein Ligases
SKP1 protein, human
Intrinsically Disordered Proteins