Macromolecular‐crowding induced protein stabilization at the residue level
0
Citation
0
Reference
10
Related Paper
Abstract:
Macromolecular crowding has significant thermodynamic and kinetic consequences for biological macromolecules. Experimental evidence has shown that crowding enhances protein association, increases the rate of folding and refolding, and affects diffusion in the cytoplasm. Statistical thermodynamic models predict that macromolecular crowding increases protein stability, yet quantitative, residue‐level experimental evidence supporting these predictions is largely absent. Here we report the first residue‐specific information about the effects of macromolecular crowding on protein stability. We used NMR‐detected amide proton exchange to quantify the effects of 300 g/L polyvinyl pyrrolidone (PVP, 40 kDa) on the stability of the globular protein chymotrypsin inhibitor 2. Crowding increases the equilibrium constant for folding by up to 100‐fold compared to dilute solution and the model of the PVP monomer. We also show that, consistent with the local unfolding model, the magnitude of the increase depends on the location of the residue in the structure. Our results demonstrate that the increase in stability upon addition of PVP is the result of macromolecular crowding.Keywords:
Macromolecular Crowding
Crowding
Globular protein
Residue (chemistry)
Macromolecular Substances
Macromolecular Crowding
Folding (DSP implementation)
Crowding
Macromolecular Substances
Protein Stability
Cite
Citations (9)
How the crowded environment inside cells affects folding, stability and structures of proteins is a vital question, since most proteins are made and function inside cells. Here we describe how crowded conditions can be created in vitro and in silico and how we have used this to probe effects on protein properties. We have found that folded forms of proteins become more compact in the presence of macromolecular crowding agents; if the protein is aspherical, the shape also changes (extent dictated by native-state stability and chemical conditions). It was also discovered that the shape of the macromolecular crowding agent modulates the folding mechanism of a protein; in addition, the extent of asphericity of the protein itself is an important factor in defining its folding speed.
Macromolecular Crowding
Folding (DSP implementation)
Crowding
Protein Stability
Cite
Citations (71)
Crowding, which characterizes the interior of all living cells, has been shown to dramatically affect biochemical processes, leading to stabilization of compact morphologies, enhanced macromolecular associations, and altered reaction rates. Due to the crowding-mediated shift in binding equilibria toward association, crowding agents were proposed to act as a metabolic buffer, significantly extending the range of intracellular conditions under which interactions occur. Crowding may, however, impose a liability because, by greatly and generally enhancing macromolecular association, it can lead to irreversible interactions. To better understand the physical determinants and physiological consequences of crowding-mediated buffering, we studied the effects of crowding, or excluded volume, on DNA structures. Results obtained from isothermal titration calorimetry (ITC) and UV melting experiments indicate that crowding-induced effects are marginal under conditions that a priori favor association of DNA strands but become progressively larger when conditions deteriorate. As such, crowding exerts "genuine" buffering activity. Unexpectedly, crowding-mediated effects are found to include enthalpy terms that favorably contribute to association processes. We propose that these enthalpy terms and preferential stabilization derive from a reconfiguration of DNA hydration that occurs in dense DNA-rich phases obtained in crowded environments.
Macromolecular Crowding
Crowding
Macromolecular Substances
Cite
Citations (93)
Macromolecular Crowding
Crowding
Folding (DSP implementation)
Biomolecule
Conformational isomerism
Macromolecular Substances
Cite
Citations (25)
Macromolecular Crowding
Crowding
Folding (DSP implementation)
Excluded volume
Macromolecular Substances
Cite
Citations (177)
Macromolecular Crowding
Co-chaperone
Chaperone (clinical)
Chemical chaperone
Mesoscopic physics
Crowding
Macromolecular Substances
Folding (DSP implementation)
Cite
Citations (48)
Macromolecular Crowding
Crowding
Folding (DSP implementation)
Excluded volume
Macromolecular Substances
Cite
Citations (41)
Macromolecular Crowding
Crowding
Brownian dynamics
Macromolecular Substances
Crowding out
Cite
Citations (94)
Based on a very simple coarse-grained colloidal model, here we implement an effective hard-sphere theory and numerical simulation to capture the general features of the association equilibria for globular proteins in crowded environment. We measure the activity coefficient, i.e., the deviation from ideal behavior of protein solution, and the crowding factor, i.e., the contribution of crowders to the association equilibria, for proteins in macromolecular crowding. The results show that the association balance in macromolecular crowding depends sensitively on the magnitude of protein–crowder attraction and the relative size of reactant to crowding agent. Since our coarse-grained model is irrelevant to the microscopic details of the molecules, it can be applied to the control of the association equilibria of many globular proteins such as bovine serum albumin, crystallin and lysozyme.
Globular protein
Macromolecular Crowding
Crowding
Macromolecular Substances
Association (psychology)
Cite
Citations (1)
Macromolecular Crowding
Chaperone (clinical)
Macromolecular Substances
Protein Stability
Co-chaperone
Cite
Citations (0)