A Next-Generation qPlus-Sensor-Based AFM Setup: Resolving Archaeal S-layer Protein Structures in Air and Liquid

TL;DR

This paper presents a next-generation AFM setup using a qPlus sensor to resolve the crystal structure of archaeal S-layer proteins in both air and liquid, demonstrating improved sample preparation and imaging techniques.

Contribution

Introduction of a new AFM protocol with advanced liquid cell and sapphire tips enabling high-resolution imaging of S-layer proteins in different environments.

Findings

Resolved S-layer crystal structure in air and liquid

Improved sample preparation for AFM imaging

Demonstrated AFM capabilities comparable to TEM in structural analysis

Abstract

Surface-layer (S-layer) proteins form the outermost envelope in many bacteria and most archaea and arrange in 2D quasi-crystalline structures via self-assembly. We investigated S-layer proteins extracted from the archaeon Pyrobaculum aerophilium with a qPlus sensor-based atomic force microscope (AFM) in both liquid and ambient conditions and compared it to transmission electron microscopy (TEM) images under vacuum conditions. For AFM scanning, a next-generation liquid cell and a new protocol for creating long and sharp sapphire tips was introduced. Initial AFM images showed only layers of residual detergent molecules (SDS), which are used to isolate the S-layer proteins from the cells. SDS was not visible in the TEM images, requiring a more thorough sample preparation for AFM measurements. These improvements allowed us to resolve the crystal-like structure of the S-layer samples with frequency-modulation AFM in both air and liquid.