Synthèses enzymatiques de néoglucoconjugués catalysées par l'alpha-glucosidase purifiée de la blatte Periplaneta americana (Linnaeus)

Enzymatic synthesis of neoglucoconjugates by purified α-glucosidase from cockroach Periplaneta americana (Linnaeus). Cockroach Periplaneta americana (Linnaeus) contains in his digestive tract an acid (pH 5,0) and mesophile (50°C) α-glucosidase. This enzyme, purified to homogeneity, hydrolyses highly maltose, sucrose and p-nitrophenyl-α-Dglucopyranoside. The ability of α-glucosidase from cockroach purified to homogeneity to catalyse transglucosylation reactions was tested using maltose and saccharose as glucosyl donors and 2-phenylethanol and phenol as acceptors. The experimental conditions were optimized in relation to the time course of the reaction, pH and concentrations of glucosyl donors and acceptors. The yields in transglucosylation reactions at 37 °C were very high and could attain 67% and 48% with 2-phenylethanol and phenol respectively as glucosyl acceptors. This α-glucosidase hydrolyzed the products formed. It seems that the products formed were the phenylethyl-α-D-glucoside and phenyl-α-D-glucoside. These results suggest that α- glucosidase from cockroach is an exoglucosidase which catalyse the splitting of the α-glucosyl residue from the non reducing terminal of the substrate to liberate α-glucose. This comportment indicates that this enzyme operated by a mechanism involving the retention of the anomeric configuration. On the basis of this work, α-glucosidase from P. americana appears to be a valuable tool for the preparation of α-neoglucoconjugates

Purification and physicochemical properties of - amylase from cockroach, Periplaneta americana (LINNAEUS), for starches saccharification

An -amylase was purified from the American cockroach Periplaneta americana (Linnaeus) to homogeneity by four steps purification via ammonium sulphate crude extract precipitation, SephacrylS-100 HR gel permeation chromatography, anion exchange chromatography on DEAE-Sepharose CL-6B and hydrophobic interaction chromatography on phenyl Sepharose CL-4B. The purification was approximatively 38.42 fold with a 24.31% yield. Optimums pH and temperature of the purified -amylase were found to be 5.6 and 55°C, respectively. The enzyme was stable up to 55°C and its pH stability was in range of 5.6 - 6.6. The KM and Vmax of the enzyme with soluble starch as substrate were 5 mg/ml and100 ìmol/min/mg, respectively, and the energy of activation (Ea), was 50.32 Kj/mol. The -amylase was inhibited by Tris, Fe3+, Ba2+, Mo+ and EDTA. While Ca2+, K+, Cu2+, Mg2+ and  para-hydroxymercuribenzoate (pHMB) activated the enzyme. Analysis of the amylolytic reaction products by HPLC showed thepresence of maltose and maltodextrin but not glucose in the starch hydrolysate (2 h of reaction). This result indicated that the amylolytic enzyme of P. americana is an -amylase (an endoamylase). Thepurified -amylase hydrolysed maltopentose, maltohexose and maltoheptose. Maltose, maltotriose and maltotetrose were not hydrolysed by this enzyme. Therefore, the purified -amylase is active only on substrates with more than four residues of glucose