Related Experiment Video
Updated: Jun 17, 2026
Identification of Hemolytic and Phospholipase Activity in Crude Extracts from Sea Anemones by Straightforward Bioassays
Published on: March 29, 2022
Molecular characterization and expression analysis of a c-type and two novel muramidase-deficient i-type lysozymes
Premruethai Supungul1, Vichien Rimphanitchayakit, Takashi Aoki
1Center of Excellence for Molecular Biology and Genomics of Shrimp, Department of Biochemistry, Faculty of Science, Chulalongkorn University, Phayathai Road, Bangkok 10330, Thailand.
Abstract:
Lysozyme is a widely distributed hydrolase possessing a hydrolytic activity against peptidoglycan in the bacterial cell wall and, hence, causing lysis of the bacteria. Two types of lysozymes; the c-type (PmLyzc) and the two catalytic residue ablated i-type lysozymes (PmLyzi1 and 2), were identified from the Penaeus monodon EST database (http://pmonodon.biotec.or.th). By RT-PCR, PmLyzc transcript was detected in all tissues: gill, antennal gland, epipodite, heart, hemocyte, hepatopancreas, eyestalk, lymphoid organ and intestine, and highly expressed in hemocyte. The expression of PmLyzi2 mRNA was highest in heart while undetected in gill, lymphoid organ and intestine. The PmLyzi1 transcript was expressed only in hepatopancreas. The up-regulation of mRNA transcription after bacterial challenge was observed only with PmLyzc. To investigate their biological activities, the three mature recombinant proteins were expressed in an Escherichia coli system. Although the turbidimetric assay revealed that only recombinant PmLyzc possessed the muramidase activity, all of them variably exhibited antimicrobial activity against both Gram-positive and -negative bacteria especially the shrimp pathogens, Vibrio species. The antimicrobial activities of recombinant PmLyzc was the most effective one. These results demonstrated that the ability of lysozyme to inhibit the growth of bacteria did not depend only on the muramidase activity. Differences in tissue expression pattern of these gene transcripts and their antimicrobial activities indicated the multifunction of lysozyme as immune defense and digestive enzymes in P. monodon.
Insights
Two Penaeus monodon lysozymes, c-type (PmLyc) and i-type (PmLyi), show varied tissue expression and antimicrobial activity. PmLyc exhibits muramidase activity and potent bacterial inhibition, suggesting multifunctional roles beyond hydrolysis in shrimp immunity.
Area of Science:
- Marine Biology
- Immunology
- Biochemistry
Background:
- Lysozyme is a key enzyme in innate immunity, hydrolyzing bacterial peptidoglycan.
- The shrimp Penaeus monodon possesses multiple lysozyme genes, including c-type and i-type.
Purpose of the Study:
- To identify and characterize c-type and i-type lysozymes from Penaeus monodon.
- To investigate the tissue-specific expression and antimicrobial activities of these lysozymes.
Main Methods:
- Expressed Sequence Tag (EST) database mining and RT-PCR for gene identification and expression analysis.
- Recombinant protein expression in Escherichia coli and biochemical assays (turbidimetric, antimicrobial).
Main Results:
- Three lysozyme transcripts (PmLyc, PmLyi1, PmLyi2) were identified with distinct tissue expression patterns.
- Only PmLyc showed muramidase activity, but all recombinant lysozymes exhibited antimicrobial effects, especially against Vibrio species.
- PmLyc demonstrated the strongest antimicrobial activity, with expression up-regulated after bacterial challenge.
Conclusions:
- Penaeus monodon lysozymes exhibit diverse tissue distribution and functional roles.
- Antimicrobial activity is not solely dependent on muramidase activity, indicating multifaceted functions in shrimp immunity and digestion.
