One‐Pot N2C/C2C/N2N Ligation To Trap Weak Protein–Protein Interactions

Weak transient protein–protein interactions (PPIs) play an essential role in cellular dynamics. However, it is challenging to obtain weak protein complexes owing to their short lifetime. Herein we present a general and facile method for trapping weak PPIs in an unbiased manner using proximity‐induced ligations. To expand the chemical ligation spectrum, we developed novel N2N (N‐terminus to N‐terminus) and C2C (C‐terminus to C‐terminus) ligation approaches. By using N2C (N‐terminus to C‐terminus), N2N, and C2C ligations in one pot, the interacting proteins were linked. The weak Ypt1:GDI interaction drove C2C ligation with t_1/2 of 4.8 min and near quantitative conversion. The Ypt1‐GDI conjugate revealed that binding of Ypt1 G‐domain causes opening of the lipid‐binding site of GDI, which can accommodate one prenyl group, giving insights into Rab membrane recycling. Moreover, we used this strategy to trap the KRas homodimer, which plays an important role in Ras signaling.