Midgut juice of Plutella xylostella strain PXR which is resistant to Cry1Ac was biochemically characterized relative to the susceptible PXS strain. The midgut juice of PXR (PXR-Juice) was shown to process Cry1Ac protoxin to 60 kDa active toxin with the same processing pattern as that of juice from PXS (PXS-Juice) in SOS-PAGE. PXS larvae which were given the Cry1Ac toxin pre-processed with PXR-Juice were killed with the same rate as that with Cry1Ac pre-activated by trypsin. PXR-Juice was found to contain three times larger amount of 66 kDa protein (P66) than PXS-Juice and the N-terminal amino acid sequence of P66 was matched to that of glucosinolate sulfatase in data base search. The protein band of P66 was coincided with the band of p-nitro phenyl sulfatase activity in zymogram. P66 purified to homogeneity in SOS-PAGE bound to Cry1Ac and soybean agglutinin, and K-D for Cry1Ac was estimated to be 718 nM with surface plasmon resonance analysis. Using purified sulfatase, K-m and V-max were estimated and involvement of the enzyme in the PXR resistance was discussed. (C) 2011 Elsevier Inc. All rights reserved.