An allosteric HTRA1-calpain 2 complex with restricted activation profile

Abstract

Significance Classic serine proteases are synthesized as inactive precursors that are proteolytically processed, resulting in irreversible activation. We report an alternative and reversible mechanism of activation that is executed by an inactive protease. This mechanism involves a protein complex between the serine protease HTRA1 and the cysteine protease calpain 2. Surprisingly, activation is restricted as it improves the proteolysis of soluble tau protein but not the dissociation and degradation of its amyloid fibrils, a task that free HTRA1 is efficiently performing. These data exemplify a challenge for protein quality control proteases in the clearing of pathogenic fibrils and suggest a potential for unexpected side effects of chemical modulators targeting PDZ or other domains located at a distance to the active site.

Rational correction of pathogenic conformational defects in HTRA1

“…In contrast, peptides that strongly activate wt HTRA1 by binding to the PDZ domain 16 had no effect on the activity of the R274Q mutant. The same effect was observed for a peptide (CAPN2.1) that activates wt HTRA1 in a PDZ-independent manner 16 (Supplementary Fig. 12a ).…”

“…15a–c ). To distinguish between these modes of action, we tested a VDAC2 derivative (VDAC2-2) which was rationally optimized for active site-binding 16 . This peptide fully activated wt HTRA1 but not R274Q (Supplementary Fig.…”

“…Future efforts should be directed to improve critical parameters of these peptides such as solubility and stability in complex biological systems. HtrA proteases are regulated by the mechanism of ligand-induced activation, which can be allosteric 8 , 16 , 33 . In bacteria, such ligands include the C-termini of β-barrel outer membrane porins serving as signals of protein folding stress 33 .…”

“…Like other HtrAs, HTRA1 displays a composite activation domain consisting of loops L1, L2, L3, and LD 1 , 13 – 15 . Activation is initiated by an interaction of the sensor loop L3 with the substrate bound to the active site or the PDZ domain 13 , 16 . Subsequently, the loop L3 interacts with loop LD of an adjacent protomer.…”

HTRA1 disaggregates α-synuclein amyloid fibrils and converts them into non-toxic and seeding incompetent species

“…HTRA1 proteolytic activity is activated by inter-monomer communication that requires trimeric assembly. Activity can be regulated through allosteric activation mediated by PDZ domain binding 30 , 31 . PDZ domains are known to mediate binding to substrates harboring β-sheets via a β-sheet augmentation mechanism, whereby the β-sheet rich region of the PDZ domain binds the β-sheet of the substrate 27 , 32 .…”