Allosteric inhibition of HTRA1 activity by a conformational lock mechanism to treat age-related macular degeneration

The trimeric serine protease HTRA1 is a genetic risk factor associated with geographic atrophy (GA), a currently untreatable form of age-related macular degeneration. Here, we describe the allosteric inhibition mechanism of HTRA1 by a clinical Fab fragment, currently being evaluated for GA treatment. Using cryo-EM, X-ray crystallography and biochemical assays we identify the exposed LoopA of HTRA1 as the sole Fab epitope, which is approximately 30 angstrom away from the active site. The cryo-EM structure of the HTRA1:Fab complex in combination with molecular dynamics simulations revealed that Fab binding to LoopA locks HTRA1 in a non-competent conformational state, incapable of supporting catalysis. Moreover, grafting the HTRA1-LoopA epitope onto HTRA2 and HTRA3 transferred the allosteric inhibition mechanism. This suggests a conserved conformational lock mechanism across the HTRA family and a critical role of LoopA for catalysis, which was supported by the reduced activity of HTRA1-3 upon LoopA deletion or perturbation. This study reveals the long-range inhibition mechanism of the clinical Fab and identifies an essential function of the exposed LoopA for activity of HTRA family proteases. The protease HTRA1 is a genetic risk factor for geographic atrophy. Here, Gerhardy et al. describe its inhibition by a clinical Fab, whose binding locks it in an inactive state. The mechanism identifies an essential function of LoopA with this protease family.

Automated summary

This study describes the allosteric inhibition mechanism of the clinical Fab fragment on HTRA1, a genetic risk factor associated with geographic atrophy. The binding of Fab to LoopA of HTRA1 locks it into an inactive state, inhibiting its catalytic activity. The study also demonstrates that the allosteric inhibition mechanism can be transferred to other proteases in the HTRA family by grafting the LoopA epitope. The findings provide insights into the inhibition mechanism of the clinical Fab and highlight the critical role of LoopA in the activity of HTRA family proteases.