RESUMO
A rhizosphere strain of Stenotrophomonas maltophilia strain MUJ that is strongly antagonistic towards fungal phytopathogens secretes to the culture medium a single form of active chitinolytic enzyme belonging to family 18 of glycosyl hydrolases. The chitinase was purified by a two-stage procedure embracing fractionation with ammonium sulfate and affinity chromatography. The molecular mass of the purified enzyme determined by SDS-PAGE was approximately 52 kDa. The enzyme demonstrated highest activity at 45°C and pH 6.8. The enzymatic protein showed considerable thermal stability during 2 h incubation at 45°C. The activity of the enzyme was strongly inhibited in the presence of Hg²âº and Cu²âº. By applying mass spectrometry analysis, the peptides derived from the purified chitinase were assigned to amino acid sequences of the type ChiA chitinases synthesized by Stenotrophomonas bacteria. The purified enzyme inhibited the growth of fungal phytopathogens belonging to the genera Fusarium, Rhizoctonia and Alternaria.
Assuntos
Proteínas de Bactérias/metabolismo , Quitinases/metabolismo , Stenotrophomonas maltophilia/enzimologia , Sequência de Aminoácidos , Antibiose , Proteínas de Bactérias/genética , Proteínas de Bactérias/isolamento & purificação , Quitina/metabolismo , Quitinases/genética , Quitinases/isolamento & purificação , Eletroforese em Gel de Poliacrilamida , Estabilidade Enzimática , Fungos/crescimento & desenvolvimento , Dados de Sequência Molecular , Peso Molecular , Stenotrophomonas maltophilia/genéticaRESUMO
Chitinolytic bacteria play an important role in degradation of chitin, one of the most abundant biopolymers in nature. These microorganisms synthesize specific enzymes, that catalyze hydrolysis of beta-1,4-glycosidic bonds in low-digestible chitin polymers, turning it into low-molecular, easy to digest compounds. During last decades many bacterial chitinolytic enzymes have been studied and characterized, mainly for their potential applications in agriculture, industry and medicine. Several chitinase classifications have been proposed, either on the base of substrate specificity or amino acid sequence similarities. X-ray crystallography and NMR spectroscopy techniques enabled the determination of three dimensional structure of some chitinases, what was helpful in explaining their catalytic mechanism. Development of biotechnology and molecular biology enables a deep research in regulation and cloning of bacterial chitinase genes.
