Characterization of Trypsin-Like Serine Protease from Lethocerus indicus Salivary Venom and its Cytotoxic Effect against Human Epidermoid Carcinoma Cell, A431
Journal Title: Biotechnology Journal International - Year 2014, Vol 4, Issue 9
Abstract
Objective: Lethocerus indicus salivary venom characterization and evaluation of extracellular degradation activity and cytotoxic effect against native human collagen type 1 and epidermoid carcinoma cell, A431. Method: Salivary venom extract was collected from adult insects by injecting 2% pilocarpine of 50 µml. Enzyme presence was detected by the apiZYM assay. The proteolytic activity was tested by the photometric and zymogram methods using specific fluorescent substrates and inhibitors. The cytotoxic activity was determined by the MTT assay and Trypan blue exclusion method. Apoptosis induction was observed using AO/EB staining solution. Digestion of extracellular matrix protein was detected against native human type I collagen. Result: L. indicus salivary venom presents amylases, proteases, carbohydrases, phosphatases and lipases. Among them, protease enzyme showed highest composition. The highest rate of proteolytic activity observed at pH 8 in 35ºC (100 %). Serine proteases present predominantly in salivary venom. Cysteine and metalloproteases are also detected. The activation energy of salivary venom is 49.86 kJ. Use of serine inhibitor, PMSF inhibited 92.77% which indicated that the maximum activity was due to serine protease. Detection of trypsin-like protease was confirmed by using PMSF and TLCK with specific substrate, BApNA. It shows significant inhibitions, 82% and 78% respectively suggesting maximum influence in salivary venom. Degradation of the fibrillar native state collagen Type I into 8 smaller peptide bands showed it importance in medical application. IC50 concentration of venom that induces cytotoxicity in epidermoid carcinoma cells, A431was 2.3 µg/ml only. It gives prominent apoptotic features such as cytoplasmic membrane blebbing, nuclear contraction, nuclear fragmentation and contact inhibition. Conclusion: We suggest that further investigation of the venom will lead to identification of active compound in L. indicus salivary venom for its potential use in therapeutic application.
Authors and Affiliations
T. Shantibala, H. Debaraj, R. K. Lokeshwari, Sarbani Giri
Keywords
Related Articles
- EP ID EP237724
- DOI 10.9734/BBJ/2014/12217
- Views 88
- Downloads 0
Stearoyl-ACP Delta 9 DesaturaseMight not be the only Factor Affecting Unsaturated Fatty Acids Concentration in Oil Palm (Elaeis Sp.)
Oil palm (Elaeis guineensis) is currently the major oil-producing plant in the world. E. oleifera is another species of oil palm that is not widely cultivated due to lower productivity. Oil extracted from E. guineensis s...
New Monoclonal Antibodies to Human IgA: Obtaining and Study of Biological Properties
Aim: Preparation of monoclonal antibodies to human IgA, investigation of their properties and selection of the most appropriate McAbs for highly sensitive and specific immunoassay tests. Methodology: Balb/c mice were use...
Listeria spp. in Raw Cow and Goat Meat in Port Harcourt, Nigeria
Aims: To determine the occurrence of Listeria monocytogenes and other Listeria spp. in raw cow and goat meat. Study Design: This work was based on a completely randomized design with two replication and the average value...
Prevalence and Antibiotic Susceptibility Pattern of Methicillin Resistant Staphylococcus aureus in Tertiary Care Hospitals
Aims: Methicillin resistant Staphylococcus aureus (MRSA) is a major nosocomial pathogen in hospitals with hospital based outbreaks world-wide. The present study was carried out to estimate the prevalence of MRSA isolates...
Field Performance and Gene Expression of Drought Stress Tolerance in Cotton (Gossypium barbadense L.)
Twenty cotton genotypes belonging to Gossypium barbadense L. were used to screen the cotton genotypes for drought stress tolerance and to study gene expression of some genes related to cotton tolerance for drought stress...