Effect of macromolecular crowding on protein binding stability: modest stabilization and significant biological consequences.
Macromolecular crowding has long been known to significantly affect protein oligomerization, and yet no direct quantitative measurements appear to have been made of its effects on the binding free energy of the elemental step of adding a single subunit. Here, we report the effects of two crowding agents on the binding free energy of two subunits in the Escherichia coli polymerase III holoenzyme. The crowding agents are found, paradoxically, to have only a modest stabilizing effect, of the order of 1 kcal/mol, on the binding of the two subunits. Systematic variations in the level of stabilization with crowder size are nevertheless observed. The data are consistent with theoretical predictions based on atomistic modeling of excluded-volume interactions with crowders. We reconcile the apparent paradox presented by our data by noting that the modest effects of crowding on elemental binding steps are cumulative, and thus lead to substantial stabilization of higher oligomers. Correspondingly, the effects of small variations in the level of crowding during the lifetime of a cell may be magnified, suggesting that crowding may play a role in increased susceptibility to protein aggregation-related diseases with aging.
Batra
Jyotica
J
Department of Physics and Institute of Molecular Biophysics, Florida State University, Tallahassee, Florida, USA.
Xu
Ke
K
Qin
Sanbo
S
Zhou
Huan-Xiang
HX
D000595:Amino Acid Sequence; D003198:Computer Simulation; D004258:DNA Polymerase III / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004259:DNA-Directed DNA Polymerase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D003911:Dextrans / Q000737:chemistry; D004926:Escherichia coli; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D005362:Ficoll / Q000737:chemistry; D005470:Fluorometry; D008956:Models, Chemical; D008957:Models, Genetic; D008958:Models, Molecular; D008969:Molecular Sequence Data; D008970:Molecular Weight; D011485:Protein Binding; D055550:Protein Stability; D017385:Sequence Homology; D013816:Thermodynamics
D003911:Dextrans; D029968:Escherichia coli Proteins; D005362:Ficoll; C505238:holE protein, E coli; D004258:DNA Polymerase III; D004259:DNA-Directed DNA Polymerase; C468316:dnaQ protein, E coli