Solubilization of Membrane Proteins using designed protein WRAPS

Mihaljevic, L.; Kim, D. E.; Eisenach, H. E.; Bandawane, P. D.; Borst, A. J.; Courbet, A.; Bettin, E.; Liu, Q.; Weidle, C.; Majumder, S.; Li, X.; Lamb, M.; Azcarraga Murray, A. N.; Ravichandran, R.; Williams, E. C.; Hu, S.; Stuart, L.; Grillova, L.; Thomson, N. R.; Chang, P.; Caimano, M. J.; Hawley, K. L.; King, N. P.; Baker, D.

doi:10.1101/2025.02.04.636539

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bioRxiv · 10.1101/2025.02.04.636539

Solubilization of Membrane Proteins using designed protein WRAPS

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Abstract

The development of therapies and vaccines targeting integral membrane proteins has been complicated by their extensive hydrophobic surfaces, which can make production and structural characterization difficult. Here we describe a general deep learning-based design approach for solubilizing native membrane proteins while preserving their sequence, fold, and function using genetically encoded de novo protein WRAPs (Water-soluble RFdiffused Amphipathic Proteins) that surround the lipid-interacting hydrophobic surfaces, rendering them stable and water-soluble without the need for detergents. We design WRAPs for both beta-barrel outer membrane and helical multi-pass transmembrane proteins, and show that the solubilized proteins retain the binding and enzymatic functions of the native targets with enhanced stability. Syphilis vaccine development has been hindered by difficulties in characterizing and producing the outer membrane protein antigens; we generated soluble versions of four Treponema pallidum outer membrane beta barrels which are potential syphilis vaccine antigens. A 4.0 [A] cryo-EM map of WRAPed TP0698 is closely consistent with the design model. WRAPs should be broadly useful for facilitating biochemical and structural characterization of integral membrane proteins, enabling therapeutic discovery by screening against purified soluble targets, and generating antigenically intact immunogens for vaccine development.

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Mihaljevic, L., Kim, D. E., Eisenach, H. E., Bandawane, P. D., Borst, A. J., Courbet, A., Bettin, E., Liu, Q., Weidle, C., Majumder, S., Li, X., Lamb, M., Azcarraga Murray, A. N., Ravichandran, R., Williams, E. C., Hu, S., Stuart, L., Grillova, L., Thomson, N. R., Chang, P., Caimano, M. J., Hawley, K. L., King, N. P., Baker, D.. 2025-02-05. Solubilization of Membrane Proteins using designed protein WRAPS. https://doi.org/10.1101/2025.02.04.636539

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