Expression, Purification and Characterization of Peptidoglycan Recognition Protein 1 From Manduca sexta (L.)

Abstract

Insects have an effective defense system against invading microbes. Peptidoglycan recognition proteins (PGRPs) in hemolymph detect peptidoglycans (a structural component of bacterial cell walls) and trigger a serine proteinase network that mediates and coordinates various host immune responses. To elucidate its biochemical functions, we have expressed Manduca sexta PGRP-1 in the baculovirus expression system and purified the protein from the conditioned culture medium. Purified PGRP-1 has a molecular mass of 19 kDa. The recombinant PGRP-1 specifically binds to Lys- and DAP-type peptidoglycans from Gram-positive and Gram-negative bacteria, respectively and leads to the proteolytic activation of prophenoloxidase. The binding affinity is high for all polymeric peptidoglycans tested, except for the one from Staphylococcus aureus. M. sexta PGRP-1 is produced at a low, constitutive level by hemocytes and fat body, and its transcripts become highly abundant in both tissues after larvae are challenged by bacterial injection.