Análise bioquímica e biológica da secreção cutânea do anuro Dermatonotus muelleri

Abstract

Poisons and secretions animals are among the most complex and rich sources of biological materials, ie new molecules with potential biotechnological applications or pharmaceutical. Accordingly, molecules isolated from amphibian skin secretions may be used as alternatives for the development of new biotechnology tool, such as those target to combat resistant pathogens or develop new industrial products. This project aimed to identify and characterize molecules in the skin secretion of Dermatonotus muelleri (only specie in the genus of the family Microhylidae), through the isolation and biochemical characterization. The secretions were filtered through 10 kDa cut-off membranes and the retained fraction was analyzed by SDS-PAGE, followed by a proteomic approach. A zymographic assay was also performed. The low molecular mass filtrate was fractionated by HPLC and analyzed by mass spectrometry and nuclear magnetic resonance. Gel electrophoresis showed protein content of various molecular masses. Proteomics analysis found relevant matches for proteins distributed in the electrophoresis gel, including proteolytic enzymes. On the other hand, the < 10kDa fraction showed to contain sugars, according to the interpretation of the NMR data. The ESI-IT-TOF analyses revealed low molecular mass molecules (<800 Da) throughout the chromatographic separation, and tryptophan could be identified among the molecules. The filtered fractions were assayed for antibiotic activity but no inhibition of bacterial growth could be observed. Interestingly, one fraction exhibited a significant increase in the growth of Staphylococcus aureus. The proteomic analysis was pertinent, demonstrating the presence of several proteins related to the animal's biology and defense mechanism. From the chemical point of view, one amino acid was detected and induced bacterial growth, which could be related to alternative mechanisms to the absence of low molecular mass antibiotic compounds.