Soluble like structure-function of β-Gal desorbed from inclusion bodies

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

Cuidad Autónoma de Buenos Aires

Congreso; Reunión Conjunta de las Sociedades de BioCiencias; 2017

SAIC, SAIB, SAI, SAA, SAB, SAFE, SAFIS, SAH, SAP

Bacterial inclusiónbodies (IBs) are mesoscopic protein aggregates commonly observed in transformedbacteria, primarily formed by recombinant proteins.Historically, IBshave been considered a hurdle for the production of soluble protein species,and many genetic and process-based strategies have been developed tominimize theirformation. However, recent research has described the use of Ibs as functionalmaterials usefull as reusable catalysts, drug delivery systems, andfunctionaltopographies in tissue engineering [1]. We have proved that β-galactosidase(β-Gal) IBs (IBsβ-Gal) can be found in an amyloidal form which holdsin non-amyloidalfunctional proteins with some particular stability properties. Also, we havedemostrated that β-Gal desorbes spontenously from IBs in low osmoticpressure media(achieve by successive dilutions). In this work we study the structure/functionrelationship of the desorbed β-Gal (β-GalD). By means of intrinsicfluorescence,infrared spectroscopy (IR) and enzymatic activity experiments we have gotevidence that β-GalD retains the structural/functional properties of thesoluble protein.Furthermore, by means of IR and DLS experiments in real time, we have foundthat while β-GalD is spontaneously released from the aggregates areorganization ofIBs occurs. This results demonstrate that the IBs isolation and the concomitantdilution steps of this sample is a simple and a proper methodology to obtainactive protein. Hence, the nature of the recombinant protein and the conditionsfor IBs formation and isolation determine the success of the desorptionof the protein in asoluble-like conformation.[1].Rinas, U., et al.,Bacterial Inclusion Bodies: Discovering Their Better Half.Trends inBiochemicalSciences, 2017.