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 naïve fat body, and the level increased 5-10-fold upon injection of the larvae with UV-killed bacteria or peptidoglycan.
Fujimoto
S
S
Department of Biochemistry and Biotechnology, Faculty of Agriculture, Tottori University, 680-8553, Tottori, Japan.
Toshimori-Tsuda
I
I
Kishimoto
K
K
Yamano
Y
Y
Morishima
I
I
D000818:Animals; D000900:Anti-Bacterial Agents / Q000097:blood / Q000302:isolation & purification; D001419:Bacteria / Q000187:drug effects; D001483:Base Sequence; D012831:Bombyx / Q000201:enzymology*; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics* / Q000302:isolation & purification; D004790:Enzyme Induction / Q000187:drug effects; D015870:Gene Expression / Q000187:drug effects; D006458:Hemolymph / Q000201:enzymology; D008969:Molecular Sequence Data; D009113:Muramidase / Q000097:blood / Q000235:genetics* / Q000302:isolation & purification*; D010457:Peptidoglycan / Q000494:pharmacology; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA
D000900:Anti-Bacterial Agents; D018076:DNA, Complementary; D010457:Peptidoglycan; D009113:Muramidase