Issue 15, 2023

A NanoCurvS platform for quantitative and multiplex analysis of curvature-sensing proteins

Abstract

The cell membrane is characterized by a rich variety of topographical features such as local protrusions or invaginations. Curvature-sensing proteins, including the Bin/Amphiphysin/Rvs (BAR) or epsin N-terminal homology (ENTH) family proteins, sense the bending sharpness and the positive/negative sign of these topographical features to induce subsequent intracellular signaling. A number of assays have been developed to study curvature-sensing properties of proteins in vitro, but it is still challenging to probe low curvature regime with the diameter of curvature from hundreds of nanometers to micrometers. It is particularly difficult to generate negative membrane curvatures with well-defined curvature values in the low curvature regime. In this work, we develop a nanostructure-based curvature sensing (NanoCurvS) platform that enables quantitative and multiplex analysis of curvature-sensitive proteins in the low curvature regime, in both negative and positive directions. We use NanoCurvS to quantitatively measure the sensing range of a negative curvature-sensing protein IRSp53 (an I-BAR protein) and a positive curvature-sensing protein FBP17 (an F-BAR protein). We find that, in cell lysates, the I-BAR domain of IRSp53 is able to sense shallow negative curvatures with the diameter-of-curvature up to 1500 nm, a range much wider than previously expected. NanoCurvS is also used to probe the autoinhibition effect of IRSp53 and the phosphorylation effect of FBP17. Therefore, the NanoCurvS platform provides a robust, multiplex, and easy-to-use tool for quantitative analysis of both positive and negative curvature-sensing proteins.

Graphical abstract: A NanoCurvS platform for quantitative and multiplex analysis of curvature-sensing proteins

Supplementary files

Article information

Article type
Paper
Submitted
11 Nov 2022
Accepted
10 Jun 2023
First published
16 Jun 2023

Biomater. Sci., 2023,11, 5205-5217

A NanoCurvS platform for quantitative and multiplex analysis of curvature-sensing proteins

C. Lu, C. Tsai, T. Jones IV, V. Chim, L. H. Klausen, W. Zhang, X. Li, Z. Jahed and B. Cui, Biomater. Sci., 2023, 11, 5205 DOI: 10.1039/D2BM01856J

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