56 research outputs found
Atomic-accuracy prediction of protein loop structures through an RNA-inspired ansatz
Consistently predicting biopolymer structure at atomic resolution from
sequence alone remains a difficult problem, even for small sub-segments of
large proteins. Such loop prediction challenges, which arise frequently in
comparative modeling and protein design, can become intractable as loop lengths
exceed 10 residues and if surrounding side-chain conformations are erased. This
article introduces a modeling strategy based on a 'stepwise ansatz', recently
developed for RNA modeling, which posits that any realistic all-atom molecular
conformation can be built up by residue-by-residue stepwise enumeration. When
harnessed to a dynamic-programming-like recursion in the Rosetta framework, the
resulting stepwise assembly (SWA) protocol enables enumerative sampling of a 12
residue loop at a significant but achievable cost of thousands of CPU-hours. In
a previously established benchmark, SWA recovers crystallographic conformations
with sub-Angstrom accuracy for 19 of 20 loops, compared to 14 of 20 by KIC
modeling with a comparable expenditure of computational power. Furthermore, SWA
gives high accuracy results on an additional set of 15 loops highlighted in the
biological literature for their irregularity or unusual length. Successes
include cis-Pro touch turns, loops that pass through tunnels of other
side-chains, and loops of lengths up to 24 residues. Remaining problem cases
are traced to inaccuracies in the Rosetta all-atom energy function. In five
additional blind tests, SWA achieves sub-Angstrom accuracy models, including
the first such success in a protein/RNA binding interface, the YbxF/kink-turn
interaction in the fourth RNA-puzzle competition. These results establish
all-atom enumeration as a systematic approach to protein structure that can
leverage high performance computing and physically realistic energy functions
to more consistently achieve atomic resolution.Comment: Identity of four-loop blind test protein and parts of figures 5 have
been omitted in this preprint to ensure confidentiality of the protein
structure prior to its public releas
Cooperative binding mitigates the high-dose hook effect
Background: The high-dose hook effect (also called prozone effect) refers to the observation that if a multivalent protein acts as a linker between two parts of a protein complex, then increasing the amount of linker protein in the mixture does not always increase the amount of fully formed complex. On the contrary, at a high enough concentration range the amount of fully formed complex actually decreases. It has been observed that allosterically regulated proteins seem less susceptible to this effect. The aim of this study was two-fold: First, to investigate the mathematical basis of how allostery mitigates the prozone effect. And second, to explore the consequences of allostery and the high-dose hook effect using the example of calmodulin, a calcium-sensing protein that regulates the switch between long-term potentiation and long-term depression in neurons. Results: We use a combinatorial model of a âperfect linker proteinâ (with infinite binding affinity) to mathematically describe the hook effect and its behaviour under allosteric conditions. We show that allosteric regulation does indeed mitigate the high-dose hook effect. We then turn to calmodulin as a real-life example of an allosteric protein. Using kinetic simulations, we show that calmodulin is indeed subject to a hook effect. We also show that this effect is stronger in the presence of the allosteric activator Ca 2+/calmodulin-dependent kinase II (CaMKII), because it reduces the overall cooperativity of the calcium-calmodulin system. It follows that, surprisingly, there are conditions where increased amounts of allosteric activator actually decrease the activity of a protein. Conclusions: We show that cooperative binding can indeed act as a protective mechanism against the hook effect. This will have implications in vivo where the extent of cooperativity of a protein can be modulated, for instance, by allosteric activators or inhibitors. This can result in counterintuitive effects of decreased activity with increased concentrations of both the allosteric protein itself and its allosteric activators. Electronic supplementary material The online version of this article (doi:10.1186/s12918-017-0447-8) contains supplementary material, which is available to authorized users
Metabolic consequences of inflammatory disruption of the blood-brain barrier in an organ-on-chip model of the human neurovascular unit
In vitro progesterone modulation on bacterial endotoxin-induced production of IL-1ÎČ, TNFα, IL-6, IL-8, IL-10, MIP-1α, and MMP-9 in pre-labor human term placenta
Quantitative Evaluation of Native Protein Folds and Assemblies by Hydrogen Deuterium Exchange Mass Spectrometry (HDX-MS)
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