Protein purification, cDNA cloning and gene expression of lysozyme from eri-silkworm, Samia cynthia ricini

Lysozyme was isolated from immunized hemolymph of Samia cynthia ricini larvae by heat treatment, cation exchange and reverse-phase chromatography. A cDNA encoding lysozyme was cloned by screening the cDNA library from immunized fat body using, as a probe, a DNA fragment obtained by PCR-based differential display method. The deduced amino acid sequence showed high homology with other chicken-type lysozymes. The calculated molecular mass of the mature peptide was 13785, which agreed precisely with that obtained by MALDI-TOF mass spectrometry of the isolated protein. The lysozyme transcripts were detected at a significant level in naive fat body, and the level increased 5-10-fold upon injection of the larvae with UV-killed bacteria or peptidoglycan. (C) 2001 Elsevier Science Inc. All rights reserved.