A Study of Glutathione S-Aryltransferase from Costelytra Zealandica

<p>An investigation has been made of the stability, purification and properties of Glutathione S-aryltransferase (Ec 2.5.1.13) from the grass-grub, Costelytra zealandica. The enzyme was found to be extremely unstable in crude homogenates of grass-grubs that had been stored frozen at -2O degrees C, but was considerably more stable in homogenates of live grass-grubs. The instability increased with increase of pH. Glutathione gave some protection against inactivation. Selective fractionation of crude homogenates with (NH4)2SO4 provided some evidence for the presence of an endogenous inhibitor of the enzyme. DEAE-cellulose chromatography and isoelectric focusing studies showed the presence of two major GSH S-aryltransferases with isoelectric points of 4.6 and 8.7. Both enzymes were present in the homogenate from a single, live, grass-grub. The molecular weight and optimum pH of each enzyme was identical within experimental error. A brief comparative study of GSH S-transferases showed the presence of GSH S-alkyl- and GSH s-alkene-transferase, but in only very small amounts compared with GSH S-aryltransferase. Differences in stability were demonstrated and some cross-specificity was indicated. Several inhibitor-substituted Sepharoses were prepared in an attempt to purify GSH s-aryltransferase by affinity chromatography. Although columns of the inhibitors removed the enzyme from solution an active enzyme could not be recovered. The effects of pH and temperature on the enzyme-catalysed reaction of GSH and 1, 2-dichloro-4-nitrobenzene (DCNB) were investigated in detail. Analysis of the variation of pKGSH with pH showed the presence of active site groups with pK approximately 9 involved in GSH binding. Calculation of the heat of ionization of these groups in the pI 8.7 enzyme, from the effect of temperature on their pK, suggested that the groups may be Lysine epsilon-NH2. Values for the enthalpy, free energy and entropy of GSH-binding to the pI 8.7 enzyme and of DCNB-binding to the enzyme-GSH complex were also obtained.</p>

A Study of Glutathione S-Aryltransferase from Costelytra Zealandica

<p>An investigation has been made of the stability, purification and properties of Glutathione S-aryltransferase (Ec 2.5.1.13) from the grass-grub, Costelytra zealandica. The enzyme was found to be extremely unstable in crude homogenates of grass-grubs that had been stored frozen at -2O degrees C, but was considerably more stable in homogenates of live grass-grubs. The instability increased with increase of pH. Glutathione gave some protection against inactivation. Selective fractionation of crude homogenates with (NH4)2SO4 provided some evidence for the presence of an endogenous inhibitor of the enzyme. DEAE-cellulose chromatography and isoelectric focusing studies showed the presence of two major GSH S-aryltransferases with isoelectric points of 4.6 and 8.7. Both enzymes were present in the homogenate from a single, live, grass-grub. The molecular weight and optimum pH of each enzyme was identical within experimental error. A brief comparative study of GSH S-transferases showed the presence of GSH S-alkyl- and GSH s-alkene-transferase, but in only very small amounts compared with GSH S-aryltransferase. Differences in stability were demonstrated and some cross-specificity was indicated. Several inhibitor-substituted Sepharoses were prepared in an attempt to purify GSH s-aryltransferase by affinity chromatography. Although columns of the inhibitors removed the enzyme from solution an active enzyme could not be recovered. The effects of pH and temperature on the enzyme-catalysed reaction of GSH and 1, 2-dichloro-4-nitrobenzene (DCNB) were investigated in detail. Analysis of the variation of pKGSH with pH showed the presence of active site groups with pK approximately 9 involved in GSH binding. Calculation of the heat of ionization of these groups in the pI 8.7 enzyme, from the effect of temperature on their pK, suggested that the groups may be Lysine epsilon-NH2. Values for the enthalpy, free energy and entropy of GSH-binding to the pI 8.7 enzyme and of DCNB-binding to the enzyme-GSH complex were also obtained.</p>

A Review on Carbon Nanotubes (CNT): Structure, Synthesis, Purification and Properties for Modern day Applications

Carbon nanotubes (CNTs), composed of graphene/graphite sheets, have been used since the 1990s and become one of the most important materials owing to its massive applications in energy, environmental and life sciences. In general, there are two types of known CNTs such as single-walled CNTs (SWCNTs) and multi-walled CNTs (MWCNTs). They have broader and novel fields of application in the fabric and textile industries, wastewater treatment, energy storage, and also as structural reinforcement materials. CNTs are reported to synthesize by arc discharge, laser ablation and chemical vapor deposition (CVD) methods where CVD technique is found to be the most versatile and largely used method. In all the methods synthesized CNTs contain various degrees of impurities which are purified by oxidation treatment, ultra-sonication, magnetic purification, etc. CNTs have wide range of industrial applications due to their remarkable mechanical, thermal, electrical, chemical and biological properties. In this review, medical and biomedical applications of CNTs and CNTs-based composites are specially focused due to their significant applications in these fields along with their structure, classification, preparation and related properties. Besides, CNTs’ toxicity and biosafety, especially on the human body, are also discussed in this review article.

Purification and properties of a phytase from Candida melibiosica 2491

Aim: To characterize the enzyme phytase produced by phytase-active Candida melibiosica 2491 for subsecuent use in feed industry. Methods: C. melibiosica 2491 had been selected among 118 strains as the most productive strain of phytase. In present study, the enzyme was first purified through electrophoresis grade in four steps: precipitation with organic solvent, ultrafiltration, gel chromatography and Denaturing gel electrophoresis (SDS–PAGE). Results: Higher levels of purification were obtained using ethanol. The gel chromatography showed an elution maximum at 11-12 fractions that characterize the corresponding one as high-molecular weight phytase. The purification level was found to be 19.5 folds with specific enzyme activity of 2.75 U/mg protein and yield – 19.64 %. Furthermore, the molecular weight of purified phytase was estimated to 35.9 кDa, with optimum of pH – at 4.5 and optimum of temperature at 55 °C. Maximum phytase activity in case of whole cells was found at 50 оС, which was less than using the purified enzyme. It was activated through 5 mM of Ba2+, 10 mM of Mn2+ and K+ ions. Total inhibition effect was achieved from Fe3+, Hg2+ and Zn2+. Copper ions (Cu2+) in concentrations at 5 mM conducted to partial inhibition effect, but at 10 mM the phytase activity was equal to zero. Low inhibition effect was determined in case of cobalt ions (Co2+) at concentrations of 10 mM. The phytase displayed broad sub­strate specificity and the Km for phytate was estimated to be 0.21 mM under the experimental conditions, while Vmax – 19.9 µМ/ml. Conclusion: Although the phytase produced by C. melibiosica 2491 is a promising enzyme to be used successfully in feed production, more investigations are needed to ensure its advantages.