Short communication: Protoplast formation from the thermophilic fungus Malbranchea sulfurea,using the thermostable chitinase and laminarinase of Paecilomyces varioti

Two thermostable enzymes produced by the thermophilic fungus Paecilomyces varioti, a chitinase and laminarinase, were used to isolate protoplasts of a thermophilic fungus, Malbranchea sulfurea. The frequency of protoplast regeneration observed (35%) was considerably higher than that obtained using commercial lytic enzymes.     

Comparative Characterization of Laminarinases from the Filamentous Marine Fungi Chaetomium Indicum Corda and Trichoderma Aureviride Rifai

Marine filamentous fungi (103 strains) isolated from various marine habitats were studied for their ability to produce extracellular O-glycosylhydrolases. Cultural filtrates of these strains were shown to contain a series of glycanases (laminarinases, amylases, cellulases, pustulanases) and glycosidases (β-glucosidases, N-acetyl-β-glucosaminidases, β-galactosidases, α-mannosidases).Two species of marine fungi from different habitats were chosen for isolation of laminarinases and detailed study on enzyme properties. The fungus Chaetomium indicum associated with the alga Fucus evanescens C. Agardh was collected near the Kuril Islands, and T. aureviride was sampled from bottom deposits of South China Sea. Properties of extracellular laminarinases were similar: temperature optimums (40–45 ^∘C), molecular masses (54–56 kDa), K _m (0.1–0.3 mg ml⁻¹). Temperature stability of laminarinase of C. indicum was significantly higher than those from T. aureveride. It is shown that these enzymes are specific to β-1,3-bonds in glucans, release predominantly glucose from laminaran and do not catalyze reaction of transglycosylation. Accoding to these data enzymes are exo-1,3-β-D-glucan-glucanohydrolases (EC 3.2.1.58). Inhibitor analysis demonstrated the significant role of tryptophan and tyrosine residues in the catalytic activity of enzymes. Molecules of T. aureviride laminarinase contained the functionally important thiol group.     

Hydrolase and glycosynthase activity of endo-1,3-beta-glucanase from the thermophile Pyrococcus furiosus

Pyrococcus furiosus laminarinase (LamA, PF0076) is an endo-glycosidase that hydrolyzes beta-1,3-glucooligosaccharides, but not beta-1,4-gluco-oligosaccharides. We studied the specificity of LamA towards small saccharides by using 4-methylumbelliferyl beta-glucosides with different linkages. Besides endo-activity, wild-type LamA has some exo-activity, and catalyzes the hydrolysis of mixed-linked oligosaccharides (Glcbeta4Glcbeta3Glcbeta-MU (Glc = glucosyl, MU = 4-methylumbelliferyl)) with both beta-1,4 and beta-1,3 specificities. The LamA mutant E170A had severely reduced hydrolytic activity, which is consistent with Glu170 being the catalytic nucleophile. The E170A mutant was active as a glycosynthase, catalyzing the condensation of alpha-laminaribiosyl fluoride to different acceptors. The best condensation yields were found at pH 6.5 and 50 degrees C, but did not exceed 30%. Depending on the acceptor, the synthase generated either a beta-1,3 or a beta-1,4 linkage.     

Chemical composition of the gastric shield of a bivalve, Zyrphea crispata, and of the teeth of the gizzard of a gastropod opisthobranch, Aplysia punctata

The chemical composition of the gastric shield of the bivalve Zyrphea crispata is very similar to that of the teeth of the gizzard of the gastropod opisthobranch Aplysia punctata. Both structures are indeed mainly made up of proteins and chitin. Their protein fractions, although heterogeneous, have almost the same amino acid composition. The proportion of free chitin is remarkably high, amounting to 74–78% of the total chitin. These cuticular structures certainly play a similar function in the gastric digestion, according to the presence of some hydrolases mainly laminarinase, the activity of which is much higher than in the crystalline style. Despite their morphological dissimilarity and specialization in two distinct classes of mollusca, the gastric shield and the teeth of the gizzard thus appear to be homologous structures on the basis of their chemical composition.     

Thermodynamics of partitioning of chitinase and laminarinase in a soya lecithin liposome system and their antifungal effect against Fusarium oxysporum

Abstract

The goal of the present work was to compare the partitioning behavior of chitinase and laminarinase (from Trichoderma spp.) in soya lecithin liposomes at different temperatures and examine the activity of the resulting microencapsulated enzymes against Fusarium oxysporum. In both cases the partition coefficients (K_o/w) were greater than 1, indicating affinity of the enzymes for microencapsulation in liposomes. Enthalpy calculations indicated that the process was endothermic in the case of laminarinase and exothermic in the case of chitinase. Soya lecithin liposomes were stable for more than 20 days. The stability of the immobilized enzyme was increased in the case of chitinase, but was not changed in the case of laminarinase. Although the antifungal effects of individual immobilized preparations decreased after microencapsulation compared with non-immobilized enzymes, they were increased by the synergistic effect of both encapsulated enzymes. The application of free or immobilized enzymes was also shown to enhance the inhibitory effect of the chemical fungicide, thiabendazole, on F. oxysporum.