Method for Mapping the Membrane Protein Nanoenvironments Using DNA Nanostructures (NanoDeep).

The distribution of proteins at the plasma membrane is not uniform, but rather many proteins localize to dynamic nanodomains. To understand the functional importance of membrane protein nanodomains, it is necessary to be able to map their composition and spatial organization en masse in cell populations. Here, we present the protocol to implement a non-microscopy-based method called NanoDeep, which enables ensemble analysis of membrane protein nanodomains. NanoDeep utilizes DNA nanoassemblies to convert information about membrane protein organization into a DNA sequencing readout. By using NanoDeep, we have previously investigated the Her2 nanoenvironments, an important membrane receptor in cancer. NanoDeep has the potential to provide a new understanding of the effects of protein composition and spatial organization on the regulation of membrane protein function. In this chapter, we describe the NanoDeep protocol as applied to Her2 nanoenvironments.