Light Activation of Protein Splicing with a Photocaged Fast Intein

Intein-mediated
protein splicing has found broad biotechnological applications. Herein,
we describe our recent result in engineering a photoactivatable intein
compatible with living mammalian cells. A photocaged cysteine amino
acid residue was genetically introduced into a highly efficient Nostoc punctiforme (Npu) DnaE intein. The resulting photocaged intein was inserted into
a red fluorescent protein (RFP) mCherry and a human Src tyrosine kinase
to create inactive chimeric proteins. A light-induced photochemical
reaction was able to reactivate the intein and trigger protein splicing.
Active mCherry and Src were formed as observed by direct fluorescence
imaging or imaging of an Src kinase sensor in mammalian cells. The
genetically encoded photocaged intein is a general optogenetic tool,
allowing effective photocontrol of primary structures and functions
of proteins.