Isaac Olusanjo Adewale

Alteration in the status of glutathione transferase of the water snail, Bulinus globosus, during aestivation and recovery.

The varying status of glutathione transferases (GSTs) in water snail, Bulinus globosus, an intermediate host of disease-causing Schistosoma haematobium (Bilharz 1852) has been investigated. The expression of GST isoenzymes in the water snail appears seasonal with about three isoenzymes appearing during raining season, when the organism is active, which may reduce to a single peak of one isoenzyme during aestivation, when the organism is inactive. GST isoenzyme is present in high concentration in all the tissues investigated namely: haemolymph, foot muscle and hepatopancreas with specific activities of 0.006 ± 0.002, 0.45 ± 0.021 and 1.33 ± 0.103 units/mg protein respectively for 1-chloro-2,4-dinitrobenzene (CDNB) as substrate. With this substrate, the specific activity of GST from the hepatopancreas appears higher than the specific activities that have been previously reported for GSTs from molluscs. Partial purification of the isoenzymes using Tris acrylic acid-based resins enabled us to observe that GST appears to be the major protein in the hepatopancreas of this organism. We also found indications for the presence of an endogenous GST inhibitor in the cytosol, whose function is yet unknown. All the traditional GST inhibitors such as cibacron blue, hematin, bromosulfophthalein and S-hexylglutathione were able to inhibit the isoenzymes effectively, with cibacron blue being the most potent. The isoenzymes however have narrow substrate specificity. We conclude that different isoenzymes of GST are expressed in the same class of molluscs, even when they belong to the same genus or species, and that the expression may depend on whether the snails are on aestivation or not.

Purification and characterization of thermostable cellulase from Enterobacter cloacae IP8 isolated from decayed plant leaf litter

Abstract Cellulases are important in the hydrolysis of lignocellulosic materials and thereby contribute to biomass conversion into fuels and chemicals. A cellulase-producing bacterium was isolated from decayed plant leaf litter in soil of a botanical garden. Based on morphological, biochemical and 16S rRNA gene sequencing, it was identified as Enterobacter cloacae IP8, with gene bank accession number NR118568.1. The bacterial cellulase was purified in a three-step procedure using lyophilization, ion exchange chromatography (QAE Sephadex A-50) and gel filtration (Biogel P-100). Two isoforms of the enzyme were purified 1.21 and 1.23 folds, respectively, with yields of 30 and 29% for isoforms A and B, respectively. Apparent molecular weights of 36.61 ± 1.40 and 14.1 ± 0.10 kDa were obtained for isoforms A and B, respectively, using gel filtration chromatography. Kinetic parameters Km and Vmax were 0.13 ± 0.04 mg/ml and 3.84 ± 0.05 U/ml/min, respectively, for isoform A and 0.58 ± 0.06 mg/ml and 13.8 ± 0.10 U/ml/min, respectively, for isoform B. Optimum pH (7.0) and temperature (60 °C) of cellulase activity were determined for both isoforms A and B. Na+ and Ca2+ enhanced the activities of both isoforms. Mg2+ inhibited the enzyme activity at concentrations 4–15 mM but, while it stimulated the activity of isoform A at concentrations 15–200 mM, it inhibited that of isoform B at same concentration range. The strong inhibition of the enzyme by ethylenediaminetetraacetic acid (EDTA) confirmed the enzyme as a metalloenzyme. These results reveal the purified cellulase from E. cloacae IP8 as a thermostable, acidic to neutral metalloenzyme, suggesting that it has good potential for biotechnological applications.