High-throughput characterization of photocrosslinker-bearing ion channel variants to map residues critical for function and pharmacology.

ABSTRACT: Incorporation of noncanonical amino acids (ncAAs) can endow proteins with novel functionalities, such as crosslinking or fluorescence. In ion channels, the function of these variants can be studied with great precision using standard electrophysiology, but this approach is typically labor intensive and low throughput. Here, we establish a high-throughput protocol to conduct functional and pharmacological investigations of ncAA-containing human acid-sensing ion channel 1a (hASIC1a) variants in transiently transfected mammalian cells. We introduce 3 different photocrosslinking ncAAs into 103 positions and assess the function of the resulting 309 variants with automated patch clamp (APC). We demonstrate that the approach is efficient and versatile, as it is amenable to assessing even complex

SUBMITTER: Braun N 

PROVIDER: S-EPMC8448361 | biostudies-literature | 2021 Sep

REPOSITORIES: biostudies-literature