A switch-on molecular biosensor for detection of caspase-3 and imaging of apoptosis of cells

Apoptosis is a form of programmed cell death that is essential for maintaining internal environmental stability.Disordered apoptosis can cause a variety of diseases;therefore,sensing apoptosis can provide help in study of mechanism of the relevant diseases and drug development.It is known that caspase-3 is a key enzyme involved in apoptosis and the expression of its activity is an indication of apoptosis.Here,we present a genetically encoded switch-on m Neon Green2-based molecular biosensor.m Neon Green2 is the brightest monomeric green fluorescent protein.The substrate of caspase-3,DEVD amino acid residues,is inserted in it,while cyclized by insertion of Nostoc punctiforme Dna E intein to abolish the fluorescence(inactive state).Caspase-3-catalyzed cleavage of DEVD linearizes m Neon Green2 and rebuilds the natural barrel structure to restore the fluorescence(activated state).The characterization exhibited that the Caspase-3 biosensor has shortened response time,higher sensitivity,and prolonged functional shelf life in detection of caspase-3 amongst the existing counterparts.We also used the Caspase-3 biosensor to evaluate the effect of several drugs on the induction of apoptosis of He La and MCF-7 tumor cells and inhibition of Zika virus invasion.