Potencial de bactérias da Antártica para produção de enzimas de interesse biotecnológico

Abstract

The Antarctic continent hosts heterogeneous regions, mainly low temperatures, high incidence of UV radiation and water deficit.In this sense, microorganisms that survive and develop in this environment possibly harbor a metabolic versatility of great biotechnological interest, including the production of a range of enzymes (esterases, lipases and chitinases, among others) adapted to cold.In this context, the objective of the present work was to select bacteria isolated from marine and terrestrial samples of Antarctica for the production of esterase, lipase and chitinase, with further emphasis on the quantification and optimization of chitinase production.For this, a collection of 560 bacteria previously isolated from diverse samples of the Antarctic continent was surveyed.The highest number of positive isolates was for esterase (n = 186, 33.21%), followed by lipase (n = 109, 19.46%) and chitinase (n = 73, 13.03%).Twenty bacteria were confirmed as chitinase-producing strains in individual tests at 5 and 15 °C up to 326 h incubation and were identified based on phylogenetic analysis of the 16S rRNA gene as Arthrobacter psychrochitiniphilus (n = 18 ), Arthrobacter cryoconiti (n = 1) and Curtobacterium luteum (n = 1), all belonging to the phylum Actinobacteria.Three chitinase-producing isolates (492, 458 and 441) were selected for further growth curve assay and quantification of total proteins (kit BCA) and enzymatic activity using 3, 5dinitrosalicylic acid (DNS) and the method using Sigma's chitin blue reagent.The best isolate was A. psychrochitiniphilus 492, which showed 10.35 U/mL of chitinase and 28.80 U/mL of reducing sugars (N-acetylglucosamine) in 80 hours of incubation.Analysis of the effect of temperature on the growth of the selected isolates showed that A. psychrochitiniphilus 492 and A. psychrochitiniphilus 441 are psychrophilic and Curtobacterium luteum 458 is a psychrotolerant mesophile.A. psychrochitiniphilus 492 was then selected for further optimization of chitinase production using the Placket-Burman (PB) experimental design, in which the effect of nine independent variables was evaluated, followed by a second planning PB, a fractionated factorial 2 5-1 , and finally a Central Composite Rotatable Desing (DCCR).The results obtained allowed to establish the optimum pH and temperature and a 7.7-fold in the chitinase production.Results gathered in this study confirmed the Antarctic environment as an arsenal of great value for the production of enzymes with different possibilities of biotechnological and industrial applications, thus reinforcing the importance of research in such polyextreme continent and the knowledge of its microbial resources.