Structural Characterization and Physiological Role of Bombyx mori Fibroinase in the Silk Gland Development

Silkworm is a highly valuable insect that produces silk
through
secretion by a silk gland. Within this gland, a type of cathepsin
L protease called Fibroinase was identified as an enzyme for hydrolyzing
the primary components of silk, including fibroin and sericin. Here,
we determined the crystal structure of Fibroinase fromBombyx mori at a resolution of 1.56 Å. Comparative
structural analysis revealed that Fibroinase adopted a similar structural
pattern with papain-type cathepsin, consisting of an N-terminal domain
and a C-terminal domain. The interface between the domains forms a
substrate-binding cleft, where the E64 inhibitor noncovalently binds
in a novel manner. Additionally, computational simulations combined
with biochemical analysis allowed us to define the binding mode and
inhibition mechanism of physiological inhibitor Bombyx cysteine protease inhibitor (BCPI) with Fibroinase. Moreover, the
expression profiles and RNA interference of Fibroinase indicated its critical role in removing silk proteins in the silk
gland lumen and the destruction of silk gland tissue during the larval–pupal
metamorphosis. These findings enhance our understanding of the structural
and biochemical features of Fibroinase and its inhibitors, while also
providing evidence for the physiological role of Fibroinase in silk
gland development.