Crystal structure of a fungal protease inhibitor from Antheraea mylitta

Abstract

Indian tasar silk is produced by a wild insect called Antheraea mylitta. Insects do not have any antigen-antibody mediated immune system like vertebrates but they produce a wide variety of effector proteins and peptides possessing potent antifungal and antibacterial activity to combat microbial attack. Antheraea mylitta expresses a fungal protease inhibitor AmFPI-1, in the hemolymph that inhibits alkaline protease of Aspergillus oryzae for protection against fungal infection. AmFPI-1 is purified from the hemolymph, crystallized and the structure is solved using the single isomorphous replacement with anomalous scattering (SIRAS) method to a resolution of 2.1 Å. AmFPI-1 is a single domain protein possessing a unique fold that consists of three helices and five β strands stabilized by a network of six disulfide bonds. The reactive site of AmFPI-1 is located in the loop formed by residues 46-66, wherein Lys54 is the P₁ residue. Superimposition of the loop with reactive sites of other canonical protease inhibitors shows that reactive site conformation of AmFPI-1 is similar to them. The structure of AmFPI-1 provides a framework for the docking of a 1:1 complex between AmFPI-1 and alkaline protease. This study addresses the structural basis of AmFPI-1's specificity towards a fungal serine protease but not to mammalian trypsin and may help in designing specific inhibitors against fungal proteases.