Isolation and characterization of a lectin-like chitinase from the hepatopancreas of freshwater prawn, Macrobrachium rosenbergii.

A lectin was isolated from the hepatopancreas of freshwater prawn, Macrobrachium rosenbergii by affinity chromatography using mucin-sepharose matrix. The purity of the isolated lectin was confirmed in native gradient PAGE that showed a single protein band of ∼37.9 kDa. In SDS-PAGE also one band of ∼43.3 kDa molecular weight was observed that indicated the protein to be a monomer. The band from the SDS-PAGE gel was identified through mass spectrometry as chitinase 1. The purified chitinase (50 μg/ml) hemagglutinated rabbit RBCs and, mucin and glucose inhibited hemagglutination with minimum concentrations of 0.1 mg/ml and 100 mM, respectively. Bacterial agglutination with Gram –ve Vibrio harveyi , Aeromonas sobria and Escherichia coli was also observed by this protein. Thus, chitinase 1 showed lectin-like properties besides its chitin hydrolytic activity. In western blot with hepatopancreas sample, rabbit antiserum against chitinase 1 cross-reacted to two additional proteins namely, chitinase 1C and obstructor E (a chitin-binding protein, CBP), besides its specific reactivity. An indirect ELISA was developed with the antiserum to quantify chitinases/CBP in hepatopancreas and serum samples of M. rosenbergii. The assay was used in samples from juvenile prawns following V. harveyi challenge. At 72 h post-challenge, significantly higher levels of chitinases/CBP were quantified in the hepatopancreas of the challenged group (1.8 ± 0.2 mg/g tissue) compared to the control (1.2 ± 0.1 mg/g tissue). This study suggests that the chitinase 1 protein with lectin-like properties is possibly induced at the protein level and can be putatively involved in the innate immune response of M. rosenbergii. • A ∼43.3 kDa Chitinase 1 was purified from the hepatopancreas of M. rosenbegii. • It possesses chitinolytic, hemagglutinating and bacterial agglutinating properties. • Antiserum to chitinase 1 cross-reacted to chitinase 1C and obstructor E proteins. • ELISA was developed to quantify chitinases/CBP in hepatopancreas and serum samples. • Vibrio harveyi challenge revealed chitinases/CBP to be inducible in nature. [ABSTRACT FROM AUTHOR]