25 research outputs found

    Chondroitinase ABC I from Proteus vulgaris: cloning, recombinant expression and active site identification

    No full text
    GalAGs (galactosaminoglycans) are one subset of the GAG (glycosaminoglycan) family of chemically heterogeneous polysaccharides that are involved in a wide range of biological processes. These complex biomacromolecules are believed to be responsible for the inhibition of nerve regeneration following injury to the central nervous system. The enzymic degradation of GAG chains in damaged nervous tissue by cABC I (chondroitinase ABC I), a broad-specificity lyase that degrades GalAGs, promotes neural recovery. In the present paper, we report the subcloning of cABC I from Proteus vulgaris, and discuss a simple methodology for the recombinant expression and purification of this enzyme. The originally expressed cABC I clone resulted in an enzyme with negligible activity against a variety of GalAG substrates. Sequencing of the cABC I clone revealed four point mutations at issue with the electron-density data of the cABC I crystal structure. Site-directed mutagenesis produced a clone with restored GalAG-degrading function. We have characterized this enzyme biochemically, including an analysis of its substrate specificity. By coupling structural inspections of cABC I and an evaluation of sequence homology against other GAG-degrading lyases, a set of amino acids was chosen for further study. Mutagenesis studies of these residues resulted in the first experimental evidence of cABC I's active site. This work will facilitate the structure–function characterization of biomedically relevant GalAGs and further the development of therapeutics for nerve regeneration

    Probiotic and prebiotic utilization in diets for free-range broiler chickens<A NAME="nt01a"></A>

    No full text
    This work aimed to evaluate the effects of probiotic associated to prebiotic on performance, carcass and cut yields, qualitative traits of meat, development and score of lesions due to coccidiosis in digestive tract of broiler chickens raised in a free-range system during 85 days. One thousand, six hundred and ninety-six day-old male broiler chicks from naked-neck ISA S757-N Label Rouge line were used in a randomized block design with factorial scheme of 4x2 and four repetitions. The effect of four additive (1 - biologic promoter + coccidiosis vaccine; 2 - biologic promoter + anti-coccidiosis; 3 - chemical promoter + coccidiosis vaccine; 4 - chemical promoter + anti-coccidiosis) and two breeding systems from 35th day of age (one with no-access to pasture or confined and the other with free-access to pasture or semi-confined) on performance parameters, carcass and cut yields, qualitative meat traits, coccidiosis development and lesions in digestive tract were evaluated. There was effect (p<0.05) of additives only on sensorial analysis (meat quality) and percentage of large intestine. Breeding systems affected (p<0.05) live weight (LW) and LW gains, being the better results observed in semi-confined birds
    corecore