RT Journal Article
SR Electronic
T1 Unfolding and identification of membrane proteins from native cell membranes
JF bioRxiv
FD Cold Spring Harbor Laboratory
SP 732933
DO 10.1101/732933
A1 Galvanetto, Nicola
A1 Maity, Sourav
A1 Ilieva, Nina
A1 Laio, Alessandro
A1 Torre, Vincent
YR 2019
UL http://biorxiv.org/content/early/2019/08/13/732933.abstract
AB Is the mechanical unfolding of proteins just a technological feat applicable only to synthetic preparations or is it applicable to real biological samples? Here, we describe all the steps necessary to deal with native membranes, from the isolation of the membrane of single cells, to the characterization and identification of the embedded membrane proteins. To do so, we combined AFM-based single-molecule force spectroscopy (SMFS) with an automatic pattern classification and the cross-matching of proteomic databases (Uniprot, PDB) to identify the unfolded proteins. We applied this method to four cell types: hippocampal and dorsal root ganglia neurons, rod outer segments and disks, and we were able to classify the unfolding of 5-10% of their total content of membrane proteins. The ability to mechanically probe proteins in their native environment enables the direct mechanical phenotyping of the membrane proteins from different cell types.