A gene (Tpen_1458) encoding a putative alpha amylase from hyperthermophilic archaeon Thermofilum pendens (TfMA) was cloned and expressed in Escherichia coli. The recombinant amylolytic enzyme was purified by Ni-NTA affinity chromatography and its catalytic properties were examined. Purified TfMA was extremely thermostable with a half-life of 60 min at an optimal temperature of 95 degrees C. TfMA activity increased to 136% in the presence of 5 mM CaCl(2). Maximal activity was measured toward gamma-cyclodextrin with a specific activity of 56 U/mg using copper bicinchoninate method. TfMA catalyzed the ring-opening reaction by cleaving one alpha-1,4-glycosidic linkage of cyclodextrin to produce corresponding single maltooligosaccharide at the initial time. The final products from cyclodextrins, linear maltooligosaccharides, and starch were glucose and maltose, and TfMA could also degrade pullulan and amylase inhibitor acarbose to panose and acarviosine-glucose, respectively. These results revealed that TfMA is a novel maltogenic amylase.
Using petroleum ether to extract an antimicrobial cerebroside from the liposoluble constituent of cervus nippon antler velvet layer was studied in this paper. Single-factor experiment was used to research the effect of liquid-to-solid ratio, standing time and centrifugation time on the extraction of crude cerebroside. The optimal extraction conditions for the cerebroside as follows: liquid-to-solid ratio 15, standing time 45 min, and centrifugation time 6 min. The extracted cerebroside was purified and identified by using silica gel column and TLC, respectively. IR analysis showed that the extraction has hydroxyl and amide groups, which was proved to be cerebroside. Meanwhile, bioactivity showed that the extraction has antibacterial activity, and the purified one is better than the crude one.
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