Levana Ferromagnetizada: uma matriz de afinidade para purificar lectina de sementes de Cratylia mollis (Cramoll 1)

AUTOR(ES)

Renata Angeli

DATA DE PUBLICAÇÃO

2006

RESUMO

Levans, fructose polymer, are composed by β-2,6 linkages and can be produced by differents species of plants and bacteria . Applications for levans have been suggested in food industries, pharmaceutics, also in medicine. Ferromagnetic particle in biomedical and industrial areas have been used. Lectins find are ubiquitously distributed in nature, and occur in plants, animals and microorganisms. Cramoll 1 is specific to glucose/mannose and was purification through 10 % (w/v) seed extract of Cratylia molli,s fractionated with ammonium sulphate and the fraction (F) 40 to 60 % of saturation (F40-60) was affinity chromatographed in Sephadex G-75 (Cramoll 1,4) followed by ion exchange chromatography in CM-cellulose (Cramoll 1 and Cramoll 4). This lectin has successfully used in different biological assays. Cramoll 3, galactose specific, was purified by the same 10 % (w/v) seed extract followed by ammonium sulphate fractionated and the fraction (F) 0 to 40% of saturation (F0-40) was ion exchange chromatographed in Sephadex G-100. Nuclear Magnetic Ressonance constituting one of the major methods for structure elucidation of biomolecules in general and this technique is used with polysaccharides. The aim of this work was to evaluate the use of Zimomonas mobilis levans insolubilized and ferromagnetic (FMZAG-12) to purify lectins fructose specifics using preparations of lectins of C. mollis seeds. Hemagglutinating activity (HA) inhibition assays were used to evaluate the specific binding of Cramoll 1, Cramoll 1,4, Cramoll 3 and Con A to levans (ZAG-12 native form and fractionated by ethanol, Z-1-81, ZAP and CP-50). Con A, Cramoll 1,4, Cramoll 3 and F40-60 were incubated to FMZAG-12 wich was used as affinity support; bound lectins were eluted with specifc carbohydrates. Electrophoresis and HA were used to analyze obtained lectins. Con A and Cramoll 1,4 bound to FMZAG-12, eluted with glucose, The eluation of Cramoll 1,4 showed its characteristic eletrophoretic pattern (two polypeptide bands). Cramoll 3 did not bind. When F40-60 was incubated with FMZAG-12, only Cramoll 1 was purified, revealing one polypeptide band (isolectin pattern). Levans inhibited differently the AH of lectins and FMZAG-12 was able to purify Cramoll 1 through a faster protocol

ASSUNTO(S)

levana lectinas de cratylia mollis levana ferromagnÃtica ciencias biologicas

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