Ribosome surface properties may impose limits on the nature of the cytoplasmic proteome

Abstract

Much of the molecular motion in the cytoplasm is diffusive, which possibly limits the tempo of processes. We studied the dependence of protein mobility on protein surface properties and ionic strength. We used surface-modified fluorescent proteins (FPs) and determined their translational diffusion coefficients (D) in the cytoplasm of Escherichia coli, Lactococcus lactis and Haloferax volcanii. We find that in E. coli D depends on the net charge and its distribution over the protein, with positive proteins diffusing up to 100-fold slower than negative ones. This effect is weaker in L. lactis and Hfx. volcanii due to electrostatic screening. The decrease in mobility is probably caused by interaction of positive FPs with ribosomes as shown in in vivo diffusion measurements and confirmed in vitro with purified ribosomes. Ribosome surface properties may thus limit the composition of the cytoplasmic proteome. This finding lays bare a paradox in the functioning of prokaryotic (endo)symbionts.

Protein interaction energy landscapes are shaped by functional and also non-functional partners

“…Interestingly, this observation holds for docked proteins corresponding to true but also 54 to arbitrary partners. Our theoretical framework enables the characterization of the energy 55 Introduction diffusion coefficients of proteins in the cytoplasm [19,27]. Positively charged proteins move 82 up to 100 times more slowly as they get caught in non-specific interactions with ribosomes 83 which are negatively charged and therefore, shape the composition of the cytoplasmic 84 proteome [27].…”

“…Most studies that aim at depicting protein interactions focus on the functional ones and on the 403 characterization of the native assembly modes [14,[47][48][49][50][51]. Nevertheless, the importance of 404 non-specific interactions and non-native assembly modes in protein interactions is no longer 405 in doubt [7,19,21,27,[52][53][54][55]. Experimental and in-silico studies showed the impact of non-406 specific interactions on the in-cell mobility of proteins [7,19,21,27].…”

“…Nevertheless, the importance of 404 non-specific interactions and non-native assembly modes in protein interactions is no longer 405 in doubt [7,19,21,27,[52][53][54][55]. Experimental and in-silico studies showed the impact of non-406 specific interactions on the in-cell mobility of proteins [7,19,21,27]. In addition, an important 407 literature describes the relationship between the physico-chemical properties of proteins and 408 their ability for non-specific interactions [7,19,21,25,53].…”

Super-resolution displacement mapping of unbound single molecules reveals nanoscale heterogeneities in intracellular diffusivity

“…Charge balance thus mandates intracellular bio(macro)molecules to take the negative charges. Whereas the backbones of DNA and RNA are known to be negatively charged, the significance of protein net charges has started to gain attention in recent years (Borgia et al, 2018;Gitlin et al, 2006;Mu et al, 2017;Schavemaker et al, 2017;Smith et al, 2016). We noticed that the most abundant proteins in the mammalian cytoplasm tend to be either strongly negatively charged or neutral (Table S2).…”

“…Intriguingly, SMdM revealed that whereas a negative net charge did not significantly affect protein diffusion, the possession of positive net charges was a key factor for diffusion slowdown, and the degree of slowdown depended on the specific subcellular environments ( Figure 4F). Interestingly, in bacteria, a recent study (Schavemaker et al, 2017) has examined the diffusion of differently charged GFP variants, and also finds that all negatively charged and neutral GFPs diffuse alike, whereas positively charged GFPs diffuse much slower, a result ascribed to interaction with ribosomes. The mammalian cell, however, is a much more complicated system.…”