Title

Single-Particle EM Reveals Extensive Conformational Variability of the Ltn1 E3 Ligase

Funding Source

National Institutes of Health (NIH)/National Center for Research Resources, NIH/National Institute of General Medical Sciences, National Institute of Neurological Disorders and Stroke, National Institute of Neurological Disorders and Stroke

Grant Number

RR017573, GM103310, R01 Grant NS075719, GM061906, RSG-11-224-01-DMC, RSG-11-224-01-DMC

Department

Department of Chemistry

Document Type

Article

Publication Date

1-29-2013

Abstract

Ltn1 is a 180-kDa E3 ubiquitin ligase that associates with ribosomes and marks certain aberrant, translationally arrested nascent polypeptide chains for proteasomal degradation. In addition to its evolutionarily conserved large size, Ltn1 is characterized by the presence of a conserved N terminus, HEAT/ARM repeats predicted to comprise the majority of the protein, and a C-terminal catalytic RING domain, although the protein's exact structure is unknown. We used numerous single-particle EM strategies to characterize Ltn1's structure based on negative stain and vitreous ice data. Two-dimensional classifications and subsequent 3D reconstructions of electron density maps show that Ltn1 has an elongated form and presents a continuum of conformational states about two flexible hinge regions, whereas its overall architecture is reminiscent of multisubunit cullin-RING ubiquitin ligase complexes. We propose a model of Ltn1 function based on its conformational variability and flexibility that describes how these features may play a role in cotranslational protein quality control.

Comments

DOI: 10.1073/pnas.1210041110

PubMed ID: 23319619

Acknowledgments

We thank Ron Milligan for review of the manuscript, Craig Yoshioka for making the data set publicly available for viewing, and Ruth Nussinov for making molecular dynamics Protein Data Bank models available. This project was supported by National Institutes of Health (NIH)/National Center for Research Resources Grant RR017573 (to C.S.P. and B.C.); NIH/National Institute of General Medical Sciences Grant GM103310 (to C.S.P. and B.C.); R01 Grant NS075719 from the National Institute of Neurological Disorders and Stroke (NINDS) of the NIH (to C.A.P.J.), and GM061906 (to S.K.D. and C.D.L.); and American Cancer Society Research Scholars Grants RSG-11-224-01-DMC (to M.D.P.) and RSG-08-298-01-TBE (to C.A.P.J.).

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