SOLUBLE LIKE STRUCTURE-FUNCTION OF beta-GAL DESORBED FROM INCLUSION BODIES

FLORES, SS; SANCHEZ, JM; NOLAN, MV; PERILLO, MA

Buenos Aires.

Otro; Reunión Conjunta de Sociedades de Biociencias; 2017

Sociedades Argentinas de Biociencias

Bacterial inclusión bodies (IBs) are mesoscopic protein aggregates commonly observed in transformed bacteria, primarily formed by recombinant proteins. Historically, IBs have been considered a hurdle for the production of soluble protein species, and many genetic and process-based strategies have been developed to minimize their formation. However, recent research has described the use of IBs as functional materials usefull as reusable catalysts, drug delivery systems, and functional topographies in tissue engineering [1]. We have proved that β-galactosidase (β-Gal) IBs (IBsβ-Gal) can be found in an amyloidal form which holds in non-amyloidal functional proteins with some particular stability properties. Also, we have demostrated that β-Gal desorbes spontenously from IBs in low osmotic pressure media (achieve by successive dilutions). In this work we study the structure/function relationship of the desorbed β-Gal (β-GalD). By means of intrinsic fluorescence, infrared spectroscopy (IR) and enzymatic activity experiments we have got evidence that β-GalD retains the structural/functional properties of the soluble protein. Furthermore, by means of IR and DLS experiments in real time, we have found that while β-GalD is spontaneously released from the aggregates a reorganization of IBs occurs. This results demonstrate that the IBs isolation and the concomitant dilution steps of this sample is a simple and a proper methodology to obtain active protein. Hence, the nature of the recombinant protein and the conditions for IBs formation and isolation determine the success of the desorption of the protein in a soluble-like conformation.1.Rinas, U., et al., Bacterial Inclusion Bodies: Discovering Their Better Half.Trends in BiochemicalSciences, 2017.