Robust and precise tools are needed to enhance the functionality
and resilience of synthetic nanoarchitectures. Here, we have employed
directed evolution and rational design to build a fast-acting molecular
superglue from a bacterial adhesion protein. We have generated the
SnoopLigase2 coupling system, a genetically encoded route for efficient
transamidation between SnoopTag2 and DogTag2 peptides. Each peptide
was selected for rapid reaction by phage display screening. The optimized
set allows more than 99% completion and is compatible with diverse
buffers, pH values, and temperatures, accelerating the reaction over
1000-fold. SnoopLigase2 directs a specific reaction in the mammalian
secretory pathway, allowing covalent display on the plasma membrane.
Transglutaminase 2 (TG2) has a network of interactions and substrates
amidst the mammalian cell surface and extracellular matrix. We expressed
a
modified TG2 with resistance to oxidative inactivation and minimal
self-reactivity. SnoopLigase2 enables TG2 functionalization with transforming
growth factor alpha (TGFα) in routes that would be impossible
through genetic fusion. The TG2:TGFα conjugate retained transamidase
activity, stably anchored TGFα for signal activation in the
extracellular environment, and reprogrammed cell behavior. This modular
toolbox should create new opportunities for molecular assembly, both
for novel biomaterials and complex cellular environments.