Fine Sampling of Sequence Space for Membrane Protein Structural Biology

• Published in: Journal of molecular biology Vol. 433; no. 15; p. 167055
• Main Authors: Loukeris, Michael, Sanghai, Zahra Assur, Vendome, Jeremie, Hendrickson, Wayne A., Kloss, Brian, Mancia, Filippo
• Format: Journal Article
• Language: English
• Published: Elsevier Ltd 23.07.2021
• Subjects: high throughput biology; integral membrane proteins; protein expression; protein purification; structural biology; structural genomics; structural genomics; TCEP-HCl; NYCOMPS; TEV; LDAO; AEBSF; protein purification; DM; high throughput biology; DDM; Ni2+-NTA; SEC; COMPPÅ; HEPES; IPTG; structural biology; integral membrane proteins; protein expression; β-OG; E. coli
• ISSN: 0022-2836; 1089-8638
• DOI: 10.1016/j.jmb.2021.167055

[Display omitted] •Membrane protein structure determination is challenging.•Membrane protein structural biology benefits from structural genomics approaches.•Good expression and detergent-stability correlate with position in sequence space.•Coarse followed by fine sampling of sequence space can increase success. We describe an enhancement of traditional genomics-based approaches to improve the success of structure determination of membrane proteins. Following a broad screen of sequence space to identify initial expression-positive targets, we employ a second step to select orthologs with closely related sequences to these hits. We demonstrate that a greater percentage of these latter targets express well and are stable in detergent, increasing the likelihood of identifying candidates that will ultimately yield structural information.