A novel bio-functional material based on mammalian cell aggresomes

Abstract

Aggresomes are protein aggregates found in mammalian cells when the intracellular protein degradation machinery is over-titred. Despite they abound in cells producing recombinant proteins of biomedical and biotechnological interest, the physiological roles of these protein clusters and the functional status of the embedded proteins remain basically unexplored. In this work, we have determined for the first time that, like in bacterial inclusion bodies, deposition of recombinant proteins into aggresomes does not imply functional inactivation. As a model , human -galactosidase A (GLA) has been expressed in mammalian cells as enzymatically active, mechanically stable aggresomes showing higher thermal stability than the soluble GLA version. Since aggresomes are easily produced and purified, we propose these particles as novel functional biomaterials with potential as carrier-free, self-immobilized catalyzers in biotechnology and biomedicine.

Functional inclusion bodies produced in the yeast Pichia pastoris

“…The value of IBs is their unusual biophysical properties that involve porosity, mechanical stability, membrane activity, protein functionality and the ability to release active protein upon internalization in mammalian cells [ 2 ]. While they are commonly observed in recombinant bacteria and mammalian cells (as functional aggresomes [ 55 , 56 ]), reports about the formation of IBs or IB-like structures in yeast cells are missing. Results obtained here indicate that IBs are indeed formed in recombinant P. pastoris upon the overproduction of an aggregation-prone model protein, under conventional production conditions and without the need to apply stress pressures such as high temperature.…”

Bacterial Inclusion Bodies: Discovering Their Better Half

“…In a recent study, a recombinant human α-galactosidase A (GLA) was produced in mammalian cells as enzymatically active, mechanically stable aggresomes [57] representing approximately 40 % of the total product. These functional aggresomes were more resistant to thermal inactivation than the soluble version of the enzyme, and were easily recovered and reused at least in five consecutive enzymatic reactions [57]. A comparative between the performance of aggresomes and IBs formed by enzymes, respective to soluble enzymes, is shown in Figure 3 C.…”

Insoluble proteins catch heterologous soluble proteins into inclusion bodies by intermolecular interaction of aggregating peptides.

“…In fact, these protein nanoclusters have been described as a source of soluble active protein obtained upon incubation in non-denaturing conditions [14][15][16] and have also been administered as biocompatible depots for tumor targeting of therapeutic proteins [17][18][19][20][21]. Furthermore, protein aggregation seems to be a common mechanism described in most of the expression systems [47][48][49] that opens up the possibility of expanding this type of strategy to proteins that are di cult to produce in prokaryotes. Therefore, the fusion of common APP to different therapeutic recombinant proteins can induce the colocalization of two recombinant proteins in IBs, obtaining protein formulations with potential synergic activities.…”