RT Journal Article
SR Electronic
T1 Chemical modification of proteins by insertion of synthetic peptides using tandem protein trans-splicing
JF bioRxiv
FD Cold Spring Harbor Laboratory
SP 2020.01.31.925495
DO 10.1101/2020.01.31.925495
A1 K.K. Khoo
A1 I. Galleano
A1 F. Gasparri
A1 R. Wieneke
A1 H. Harms
A1 M.H. Poulsen
A1 H.C. Chua
A1 M. Wulf
A1 R. Tampé
A1 S.A. Pless
YR 2020
UL http://biorxiv.org/content/early/2020/02/02/2020.01.31.925495.abstract
AB Manipulation of proteins by chemical modification is a powerful way to decipher their function or harness that function for therapeutic purposes. Despite recent progress in ribosome-dependent and semi-synthetic chemical modifications, these techniques sometimes have limitations in the number and type of modifications that can be simultaneously introduced or their application in live eukaryotic cells. Here we present a new approach to incorporate single or multiple post-translational modifications or non-canonical amino acids into soluble and membrane proteins expressed in eukaryotic cells. We insert synthetic peptides into proteins of interest via tandem protein trans-splicing using two orthogonal split intein pairs and validate our approach by investigating different aspects of GFP, NaV1.5 and P2X2 receptor function. Because the approach can introduce virtually any chemical modification into both intracellular and extracellular regions of target proteins, we anticipate that it will overcome some of the drawbacks of other semi-synthetic or ribosome-dependent methods to engineer proteins.