Fluitt A, Pienaar E, Viljoen H. Ribosome kinetics and aa-tRNA competition determine rate and fidelity of peptide synthesis. Comput Biol Chem. 2007;31(5–6):335–46. The interference of protein corona with cytotoxicity assays (including the 3-[4,5-dimethylthiazol-2yl]-2,5-diphenyltetrazolium bromide (MTT) assay) was one of the first issues raised 110. The formation of the protein corona can substantially change the composition and nutritional balance of cell culture media in static settings mainly because of the attraction of proteins, amino acids and vitamins to the surface of NPs and because of variations in protein conformation after interactions with the surface of NPs. As a result, the modified cell culture medium itself can induce cytotoxic effects under static in vitro conditions and may cause errors in NP toxicity outcomes. Two main strategies can be used to achieve reliable NP toxicological data. The first approach is to use a bioreactor, in which the use of dynamic flow can minimize the protein–NP adsorptive effects in cell culture media, thereby avoiding NP-induced variabilities in media composition. The second strategy is to introduce NPs to the cell culture media first and incubate them for an hour (to ensure that equilibrium among the biomolecules on the NP surface is reached) and then collect the particles and introduce them to cells in fresh culture media to minimize the effect of the protein corona on the composition of the culture media. Natoli, G. Maintaining cell identity through global control of genomic organization. Immunity 33, 12–24 (2010).

Mullen, A.C. et al. Master transcription factors determine cell-type-specific responses to TGF-β signaling. Cell 147, 565–576 (2011). Moszer I, Glaser P, Danchin A. SubtiList: a relational database for the Bacillus subtilis genome. Microbiology. 1995;141:261–8. Sambrook J, Fritsch EF, Maniatis T. Molecular cloning: a laboratory manual. Cold Spring Harbor: Cold Spring Harbor Laboratory Press; 1989. Kasana RC, Salwan R, Yadav SK. Microbial proteases: detection, production, and genetic improvement. Crit Rev Microbiol. 2011;37:262–76.Stengel F, Aebersold R, Robinson CV (2012) Joining forces: integrating proteomics and cross-linking with the mass spectrometry of intact complexes. Mol Cell Proteomics 11(R111):014027 Liu H, Wang X, Yang S, Wang R, Wang T. Saturation mutagenesis and self-inducible expression of trehalose synthase in Bacillus subtilis. Biotechnol Prog. 2019;35:e2826. Clarke, S.L. et al. Human developmental enhancers conserved between deuterostomes and protostomes. PLoS Genet. 8, e1002852 (2012). By studying the spatial organization of the protein corona on polystyrene NPs, the location of adsorbed proteins was discovered to be randomly distributed. Yet, these randomly adsorbed proteins possessed functional epitopes that could bind to receptors 49. Similarly, adsorbed lipoproteins on 2D graphene flakes were found to possess several functional epitopes capable of binding to receptors in the liver 50. Furthermore, analysis of adsorbed low-density lipoproteins and IgG on SiO 2 NPs revealed functional epitopes, which enhanced NP uptake in human lung and embryonic kidney cells 51. Overall, the functionality of the protein corona, which is influenced by nanomaterial physiochemical properties, protein varieties and concentrations in biofluids and ambient parameters (such as pH and temperature), allows for the specific recognition of cell receptors and suggests that there may exist multivalent interactions between cells and NPs mediated by rare corona proteins that are, to date, not well understood.

Schindelin J, Arganda-Carreras I, Frise E, Kaynig V, Longair M, Pietzsch T, Preibisch S, Rueden C, Saalfeld S, Schmid B. Fiji: an open-source platform for biological-image analysis. Nat Methods. 2012;9:676–82. Assuming that translation elongation is limited by the transport of ternary complexes towards the actively translating ribosomes, we established a model that describes diffusive motion of ternary complexes in a three-dimensional space. Furthermore, collisions of ternary complexes with tRNA-free ribosomes were defined as successful encounters resulting in instantaneous reactions. To minimize computational efforts, the reaction volume was set to 0.064 µm 3 for preliminary studies, and time steps t enc,s between two successful encounters were evaluated.Bruce JE (2012) In vivo protein complex topologies: sights through a cross-linking lens. Proteomics 12:1565–1575