Molecular cloning, recombinant protein expression, tissue distribution and functional analysis of a new c-type lysozyme from Lezhi black goat rumen

Three major distinct types of lysozymes have been identified in the animal kingdom and most lysozymes cloned from ruminants belong to the chicken-type (c-type). In this study, a new c-type lysozyme gene, named LZRLyz, was cloned and sequenced from the Lezhi black goat rumen. The LZRLyz cDNA has a 444 bp open reading frame (ORF) encoding a 147 amino acid polypeptide. The encoded polypeptide is predicted to have an 18 amino acid signal peptide, and a 129 amino acid mature protein with an isoelectric point (pI) of 6.08. The LZRLyz amino acid sequence shares 70.27% identity with the Capra hircus blood lysozyme and is grouped with other ruminants c-type lysozymes using the phylogenetic tree estimated by Neighbor-Jointing method. The recombinant expressed LZRLyz protein (pET-rLZR) shows a molecular mass of similar to 33 kDa, which is consistent with the predicted fusion protein molecular mass and shows antimicrobial activity. Quantitative real-time RT-PCR analyses revealed that LZRLyz transcripts are expressed in all tested tissues with the predominant expression being observed in rumen and the weakest one in spleen. Results of this study suggest that the LZRLyz gene represents a new c-type lysozyme gene that likely functions in Lezhi black goat host immunity and digestive systems.