One-shot design elevates functional expression levels of a voltage-gated potassium channel

Jonathan Jacob Weinstein, Chandamita Saikia, Izhar Karbat, Adi Goldenzweig, Eitan Reuveny, Sarel Jacob Fleishman

Research output: Contribution to journalArticlepeer-review

Abstract

Membrane proteins play critical physiological roles as receptors, channels, pumps, and transporters. Despite their importance, however, low expression levels often hamper the experimental characterization of membrane proteins. We present an automated and web-accessible design algorithm called mPROSS (https://mPROSS.weizmann.ac.il), which uses phylogenetic analysis and an atomistic potential, including an empirical lipophilicity scale, to improve native-state energy. As a stringent test, we apply mPROSS to the Kv1.2–Kv2.1 paddle chimera voltage-gated potassium channel. Four designs, encoding 9–26 mutations relative to the parental channel, were functional and maintained potassium-selective permeation and voltage dependence in Xenopus oocytes with up to 14-fold increase in whole-cell current densities. Additionally, single-channel recordings reveal no significant change in the channel-opening probability nor in unitary conductance, indicating that functional expression levels increase without impacting the activity profile of individual channels. Our results suggest that the expression levels of other dynamic channels and receptors may be enhanced through one-shot design calculations.

Original languageEnglish
Article numbere4995
JournalProtein Science
Volume33
Issue number6
Early online date15 May 2024
DOIs
StatePublished - Jun 2024

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology

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