AN ERP60-LIKE PROTEIN FROM THE FILARIAL PARASITE DIROFILARIA-IMMITIS HAS BOTH TRANSGLUTAMINASE AND PROTEIN DISULFIDE-ISOMERASE ACTIVITY

Transglutaminases (TGases; EC 2.3.2.13) are a family of enzymes that c atalyze calcium-dependent covalent cross-linking of cellular proteins by establishing epsilon-(gamma-glutamyl)lysine isopeptide bonds, These covalent isopeptide bonds are of great physiological significance bec ause they are highly resistant to proteolysis, denaturants, and reduci ng agents, Prior studies have demonstrated the presence of isopeptide bonds in the sheath and cuticle of filarial parasites, suggesting an i mportant role for TGase-catalyzed reactions during the growth and deve lopment of filarial nematodes, Herein we report the identification and cloning of a cDNA encoding a TGase from the dog heartworm Dirofilaria immitis (DiTG), The DiTG expressed in Escherichia coil (recombinant D iTG) was able to catalyze calcium-dependent cross-linking reactions, T he derived amino acid sequence of the DiTG cDNA (pDiTG) predicts a pro tein of 57.1 kDa and includes an N-terminal hydrophobic signal peptide , The pDiTG has no sequence similarity with any of the known TGases, b ut it has significant homology to protein disulfide isomerase (PDI) an d, particularly, to the PDI-related endoplasmic reticulum protein ERp6 0, a PDI isoform found in the lumen of endoplasmic reticulum, As predi cted from the amino acid sequence homology, recombinant DiTG catalyzed the isomerization of intramolecular disulfide/sulfhydryl bonds in den atured RNase in vitro as effectively as did mammalian PDI, Conversely, purified PDI from bovine liver could catalyze protein cross-linking r eactions in a Ca2+-dependent manner. This report describes the dual ca talytic activity of TGase and PDI in post- and/or cotranslational modi fication of newly synthesized proteins, These TGase-catalyzed posttran slational modifications may play a pivotal role in the synthesis of ne w cuticle during the growth and maturation of filarial parasites.