Mosquitocidal activity of thermostable extra cellular glycoprotein of Pseudomonas fluorescens Migula
Usharani Brammacharry, Muthuraj Muthaiah
The present study describes the thermostability of glycoprotein of extra cellular mosquitocidal proteins of Pseudomonas fluorescens Migula and its involvement during pathogenesis in insects against larvae and pupae of Culex quinquefasciatus. Fluorescent glycoprotein detection assay was employed for the detection of glycoprotein. The upper and lower confidential limits of LC 50 and LC 90 values obtained through Probit regression analysis. The amino sugars like D-maltose, D-mannose, D-xylose and sialic acid were checked and determined in purified mosquitocidal protein. Two mosquitocidal proteins corresponding to 55 kDa and 35 kDa mosquitocidal proteins on 10% SDS-PAGE indicated that these proteins could withstand and were stable even at an elevated temperature of 121°C for 15 minutes. The LC50 value of crude and pure protein was 0.3 ug/ml and 0.03 ug/ml for both pre- and post-treatment. P value of liner regression analysis is <0.0001 at 95% confidence intervals, the results of determination of amino sugars D-maltose, D-mannose, D-xylose test is statistically significant. The FT IR spectra of mosquitocidal protein showed amide bonds, Carbohydrates and N-acetyl amino sugars, aliphatic hydrocarbons and hydroxyl groups. Hence, the presence of N-acetyl amino sugar moiety in the mosquitocidal protein confirmed it was glycoprotein. The 0.155 µmol/mg N-acetyl Neuraminic acid was estimated and its presence in the active mosquitocidal protein confirmed it was glycoprotein. The peak at 1.5, 2.4, and 3.3 to 3.5 ppm of 13 C NMR spectrums strongly confirmed the presence of N-acetyl amino sugars. It is concluded that the novel extra cellular mosquitocidal protein of Pseudomonas fluorescens Migula is thermostable glycoprotein.