Post-production protein stability: trouble beyond the cell factory

Abstract

Being protein function a conformation-dependent issue, avoiding aggregation during production is a major challenge in biotechnological processes, what is often successfully addressed by convenient upstream, midstream or downstream approaches. Even when obtained in soluble forms, proteins tend to aggregate, especially if stored and manipulated at high concentrations, as is the case of protein drugs for human therapy. Post-production protein aggregation is then a major concern in the pharmaceutical industry, as protein stability, pharmacokinetics, bioavailability, immunogenicity and side effects are largely dependent on the extent of aggregates formation. Apart from acting at the formulation level, the recombinant nature of protein drugs allows intervening at upstream stages through protein engineering, to produce analogue protein versions with higher stability and enhanced therapeutic values.

Intracellular CXCR4+ cell targeting with T22-empowered protein-only nanoparticles

“…These features are suggestive of proteolytic stability, tight architecture, and regular organization of T22-GFP-H6 building blocks, in contrast with the random particle size and morphologies observed during amorphous protein aggregation, even in the form of soluble aggregates. [24][25][26][27] Indeed, the mean size of T22-GFP-H6, measured by dynamic light scattering, was 13.45 nm (Figure 6C), a value fully compatible with images of these nanoparticles under transmission electron microscopy ( Figure 6D) showing relatively monodispersed entities. To assess further the structural and functional stability of T22-empowered nanoparticles, we determined their size distribution after storage for one year at -80°C, and also at room temperature for an additional 24 hours, followed by one additional step of freezing and thawing.…”

Biocatalytic potential of laccase-like multicopper oxidases from Aspergillus niger

“…In this sense, it would be interesting to test its suitability for applications related to the food industry, as A. niger is a safe production organism and many of its enzymes are considered generally recognized as safe (GRAS) by the United States Food and Drug Administration [47]. Also, the possibility to use industrial A. niger strains to overproduce the native form of McoB could overcome issues related to production yields and stability [48], that can occur during heterologous expression of recombinant proteins, and are less expected in homologous expression systems [6]. Interestingly, the production yields of this enzyme have been recently optimized, together with those of other A. niger MCOs [49].…”

A Nanostructured Bacterial Bioscaffold for The Sustained Bottom-Up Delivery of Protein Drugs

“…This is simultaneously achieved by the natural adhesiveness of IBs, which promotes strong cell attachment, combined with pERK-mediated stimulation of cell division through topographical modification, subsequent cell-sensing and mechanotransduction events [21]. The properties of IBs as self-organizing microbial materials have been extensively discussed elsewhere [13,17,[22][23][24][25][26][27][28]. Since they contain important amounts of functional proteins [17,29,30], and in addition, show an unexpected cell membrane penetrability [31][32][33], IBs rapidly enter exposed mammalian cells and release significant amounts of the forming protein, namely, its natural building block.…”