Single-molecule study on the orientation of coiled-coil proteins attached to gold nanostructures

TL;DR

This study investigates the orientation and arrangement of coiled-coil proteins on gold nanostructures, demonstrating a method for controlled protein patterning at the single-molecule level using amino acid-specific attachment.

Contribution

It introduces a novel approach to control and analyze protein orientation on nanostructures via two-point attachment using natural amino acids.

Findings

Proteins can be aligned between nanostructure sidewalls.

Varying cysteine residues affects protein structure and attachment.

Molecular dynamics help interpret protein positional distributions.

Abstract

Methods for patterning biomolecules on a substrate at the single molecule level have been studied as a route to sensors with single-molecular sensitivity or as a way to probe biological phenomena at the single-molecule level. However, the arrangement and orientation of single biomolecules on substrates has been less investigated. Here, we examined the arrangement and orientation of two rod-like coiled-coil proteins, cortexillin and tropomyosin, around patterned gold nanostructures. The high aspect ratio of the coiled coils made it possible to study their orientations and to pursue a strategy of protein orientation via two-point attachment. The proteins were anchored to the surfaces using thiol groups, and the number of cysteine residues in tropomyosin was varied to test how this variation affected the structure and arrangement of the surface-attached proteins. Molecular dynamics studies were used to interpret the observed positional distributions. Based on initial studies of protein attachment to gold post structures, two 31-nm-long tropomyosin molecules were aligned between the two sidewalls of a trench with a width of 68 nm. Because the approach presented in this study uses one of twenty natural amino acids, this method provides a convenient way to pattern biomolecules on substrates using standard chemistry.