Poster - ?Euglena gracilis expansin: production and characterization of its binding to insoluble substrates?

CALLONI, RD; MUCHUT, RJ; ARIAS, DG; IGLESIAS, AA; GUERRERO, SA

Paraná

Congreso; SAIB LIV Reunión Anual; 2018

SAIB

Euglena gracilis is a fresh water protist with a large metabolic capacity because it is able to grow photosynthetically or heterotrophically. It is a source of products of interest such as wax esters, vitamins, amino acids and polyunsaturated fatty acids. We have identified in a transcriptomic work made in E. gracilis a transcript that codes for an expansin (EgExp). These proteins are relatively small (~ 25 kDa) with a ―loosening‖ effecton cellulosic network lacking of hydrolytic action. They have been already found in plants and plant-pathogens such as bacteria and fungi. E. gracilis does not have a carbohydrate-based cell wall and is not reported to be a plant pathogen. However, it produces an insoluble and crystalline β?1,3 glucan called paramylon as storage polymer. Thus, the function of EgExp remains unknown in this microorganism.EgExp was expressedin Escherichia coli BL21 (DE3) cells as inclusion bodies. A solubilization protocol with urea 8 M was applied to denaturing the insoluble protein. After that, the chaotropic agent was removed through by dialysis. In order to characterize this protein, we evaluated the binding capacity to insoluble substrates. We observed it can bind to filter paper, sawdust and chitin, but not to paramylon. We performed the Langmuir isotherms as previously described to determine that the maximum binding capacity of EgExp is similar to that reported for other bacterial expansins. This result supports the right assignation structure- function to the polypeptide being necessary further work to in depth analyze the functionality ofEgExp in E. gracilis.