Molecular chaperones maximize the native state yield per unit time by driving substrates out of equilibrium

Molecular chaperones have evolved to facilitate folding of proteins and RNA in vivo where spontaneous self-assembly is sometimes prohibited. Folding of Tetrahymena ribozyme, assisted by the RNA chaperone CYT-19, surprisingly shows that at physiological Mg2+ ion concentrations, increasing the chaperone concentration reduces the yield of native ribozymes. In contrast, the more extensively investigated protein chaperone GroEL works in exactly the opposite manner—the yield of native substrate increases with the increase in chaperone concentration. Thus, the puzzling observation on the assisted-ribozyme folding seems to contradict the expectation that a molecular chaperone acts as an efficient annealing machine. We suggest a resolution to this apparently paradoxical behavior by developing a minimal stochastic model that captures the essence of the Iterative Annealing Mechanism (IAM), providing a unified description of chaperone mediated-folding of proteins and RNA. Our theory provides a general relation involving the kinetic rates of the system, which quantitatively predicts how the yield of native state depends on chaperone concentration. By carefully analyzing a host of experimental data on Tetrahymena (and its mutants) as well as the protein Rubisco and Malate Dehydrogenase, we show that although the absolute yield of native states decreases in the ribozyme, the rate of native state production increases in both the cases. By utilizing energy from ATP hydrolysis, both CYT-19 and GroEL drive their substrate concentrations far out of equilibrium, in an endeavor to maximize the native yield in a short time. Our findings are consistent with the general expectation that proteins or RNA need to be folded by the cellular machinery on biologically relevant timescales, even if the final yield is lower than what equilibrium thermodynamics would dictate. Besides establishing the IAM as the basis for functions of RNA and protein chaperones, our work shows that cellular copy numbers have been adjusted to optimize the rate of native state production of the folded states of RNA and proteins under physiological conditions.Significance statementMolecular chaperones have evolved to assist the folding of proteins and RNA, thus avoiding the deleterious consequences of misfolding. Thus, it is expected that increasing chaperone concentration should lead to an enhancement in native yield. While this has been observed in GroEL-mediated protein folding, experiments on Tetrahymena ribozyme folding assisted by CYT-19, surprisingly show the opposite trend. Here, we reconcile these divergent experimental observations by developing a unified stochastic model of chaperone assisted protein and RNA folding. We show that chaperones drive their substrates out of equilibrium, and in the process maximize the rate of native substrate production rather than the absolute yield or the folding rate. In vivo the number of chaperones is regulated to optimize their functions.