Characterization of an A3G-VifHIV-1-CRL5-CBFβ Structure Using a Cross-linking Mass Spectrometry Pipeline for Integrative Modeling of Host-Pathogen Complexes

Robyn M Kaake; Ignacia Echeverria; Seung Joong Kim; John Von Dollen; Nicholas M Chesarino; Yuqing Feng; Clinton Yu; Hai Ta; Linda Chelico; Lan Huang; John Gross; Andrej Sali; Nevan J Krogan

doi:10.1016/j.mcpro.2021.100132

Characterization of an A3G-Vif_HIV-1-CRL5-CBFβ Structure Using a Cross-linking Mass Spectrometry Pipeline for Integrative Modeling of Host-Pathogen Complexes

Mol Cell Proteomics. 2021:20:100132. doi: 10.1016/j.mcpro.2021.100132. Epub 2021 Aug 11.

Authors

Robyn M Kaake¹, Ignacia Echeverria², Seung Joong Kim³, John Von Dollen⁴, Nicholas M Chesarino⁵, Yuqing Feng⁶, Clinton Yu⁷, Hai Ta⁸, Linda Chelico⁶, Lan Huang⁷, John Gross⁹, Andrej Sali¹⁰, Nevan J Krogan¹¹

Affiliations

¹ Department of Cellular and Molecular Pharmacology, California Institute for Quantitative Biosciences, University of California, San Francisco, San Francisco, California, USA; Quantitative Biosciences Institute, University of California, San Francisco, San Francisco, California, USA; Gladstone Institute of Data Science and Biotechnology, J. David Gladstone Institutes, San Francisco, California, USA.
² Department of Cellular and Molecular Pharmacology, California Institute for Quantitative Biosciences, University of California, San Francisco, San Francisco, California, USA; Department of Bioengineering and Therapeutic Sciences, University of California, San Francisco, San Francisco, California, USA.
³ Department of Bioengineering and Therapeutic Sciences, University of California, San Francisco, San Francisco, California, USA.
⁴ Department of Cellular and Molecular Pharmacology, California Institute for Quantitative Biosciences, University of California, San Francisco, San Francisco, California, USA; Quantitative Biosciences Institute, University of California, San Francisco, San Francisco, California, USA.
⁵ Divisions of Human Biology and Basic Sciences, Fred Hutchinson Cancer Research Center, Seattle, Washington, USA.
⁶ Department of Biochemistry, Microbiology, Immunology, University of Saskatchewan, Saskatoon, Saskatchewan, Canada.
⁷ Department of Physiology & Biophysics, University of California, Irvine, California, USA.
⁸ Department of Pharmaceutical Chemistry, University of California, San Francisco, San Francisco, California, USA.
⁹ Quantitative Biosciences Institute, University of California, San Francisco, San Francisco, California, USA; Department of Pharmaceutical Chemistry, University of California, San Francisco, San Francisco, California, USA.
¹⁰ Quantitative Biosciences Institute, University of California, San Francisco, San Francisco, California, USA; Department of Bioengineering and Therapeutic Sciences, University of California, San Francisco, San Francisco, California, USA; Department of Pharmaceutical Chemistry, University of California, San Francisco, San Francisco, California, USA. Electronic address: sali@salilab.org.
¹¹ Department of Cellular and Molecular Pharmacology, California Institute for Quantitative Biosciences, University of California, San Francisco, San Francisco, California, USA; Quantitative Biosciences Institute, University of California, San Francisco, San Francisco, California, USA; Gladstone Institute of Data Science and Biotechnology, J. David Gladstone Institutes, San Francisco, California, USA. Electronic address: nevan.krogan@ucsf.edu.

Abstract

Structural analysis of host-pathogen protein complexes remains challenging, largely due to their structural heterogeneity. Here, we describe a pipeline for the structural characterization of these complexes using integrative structure modeling based on chemical cross-links and residue-protein contacts inferred from mutagenesis studies. We used this approach on the HIV-1 Vif protein bound to restriction factor APOBEC3G (A3G), the Cullin-5 E3 ring ligase (CRL5), and the cellular transcription factor Core Binding Factor Beta (CBFβ) to determine the structure of the (A3G-Vif-CRL5-CBFβ) complex. Using the MS-cleavable DSSO cross-linker to obtain a set of 132 cross-links within this reconstituted complex along with the atomic structures of the subunits and mutagenesis data, we computed an integrative structure model of the heptameric A3G-Vif-CRL5-CBFβ complex. The structure, which was validated using a series of tests, reveals that A3G is bound to Vif mostly through its N-terminal domain. Moreover, the model ensemble quantifies the dynamic heterogeneity of the A3G C-terminal domain and Cul5 positions. Finally, the model was used to rationalize previous structural, mutagenesis and functional data not used for modeling, including information related to the A3G-bound and unbound structures as well as mapping functional mutations to the A3G-Vif interface. The experimental and computational approach described here is generally applicable to other challenging host-pathogen protein complexes.

Keywords: architectures of host–pathogen complexes; cross-linking mass spectrometry; integrative structure modeling.

Publication types

Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't

MeSH terms

APOBEC-3G Deaminase / chemistry*
Core Binding Factor beta Subunit / chemistry*
Cullin Proteins / chemistry*
Mass Spectrometry
Models, Molecular
Ubiquitin-Protein Ligases / chemistry*
vif Gene Products, Human Immunodeficiency Virus / chemistry*

Substances

Core Binding Factor beta Subunit
Cullin Proteins
vif Gene Products, Human Immunodeficiency Virus
vif protein, Human immunodeficiency virus 1
Ubiquitin-Protein Ligases
APOBEC-3G Deaminase

Abstract

Publication types

MeSH terms

Substances

Grants and funding