Proteins can mistakenly use Lassos itself to help explain abnormal modification behaviors

Proteins are long molecules that need to fold into complex three-dimensional structures to perform cellular functions. This folding process can sometimes lead to misfolded proteins, which can potentially lead to disease if not corrected. Now, new research explains potential mechanisms that can help explain why some proteins redeploy in different patterns than expected.

Researchers led by a Pennsylvania chemist discovered that proteins can incorrectly intertwin segments and can create barriers to the normal folding process. Correcting this misfold requires high energy or widespread deployment, slowing down the folding process, leading to unexpected patterns first observed in the 1990s.

“A misfolded protein can malfunction and lead to illness,” said Ed O’Brien, professor of chemistry at the Eberly Science of Science, a co-employed by the Pennsylvania Institute of Computational and Data Science and leader of the research team. “So understanding the mechanisms involved in the folding process could help researchers prevent or develop treatment for diseases caused by accidentally folding.”

A paper describing a study that uses a combination of computer simulations and refolding experiments to explain the folding rate of a protein called phosphoglycerate kinase (PGK) appeared today (March 14th) in the Journal of Science Advancements.

“For most proteins, we model the folding process as if there were two folded or expanded states,” says Yang Jiang, professor of chemistry research at Every University of Science in Pennsylvania and the first author of the paper.

“Tracking the progression of proteins from expansion to folding gives us a distinctive time-dependent pattern called protein folding. Usually, the percentage of expanded proteins is experiencing until essentially all proteins are folded, but some proteins do not fit this pattern.

Anomalous folding patterns of PGK were first observed experimentally over 25 years ago. Most proteins fit the “two-state” model of exponential folding rates, but the molecules of PGK reached a fully folded state according to different patterns. This new pattern was described as “repeated dynamics of extended extensibility,” but the structural mechanisms that explained this difference have remained a mystery until now.

The researchers hypothesized that a class of misfolded folds in the recently described class could be involved in PGK deviations from the traditional two-state folding model.

“Non-covalent lassoentanglement is a class of misfolding that recently identifies where a protein loop traps another segment of the protein, essentially trapping a place where it is incorrectly intertwined,” O’Brien said. “It helps explain why we see stretchy extension kinetics when proteins like PGK are prone to misfolding of this type.”

To test this hypothesis, the researchers first built a computer model to simulate the folding process of PGK. Their simulations replicated the kinetics of elongation velocity seen in previous experiments. We then investigated the intermediate stages of the simulation folding process to see if there were structural changes that could explain the extended redeployment.

“We found some examples of misfolding involving entanglement,” Jiang said. “Sometimes, new entanglements formed, sometimes not entanglements, which are part of the protein’s native structure. The simulations removed these misfolding events and found that the protein was folded in a typical two-state exponential pattern.”

To confirm the results of their simulations, the research team, including Johns Hopkins University experimenter Stephen Fried and lab members, examined structural variation in PGK during the experimental remodeling. They found that the erroneously folded state predicted in the simulation coincided with structural signals observed experimentally with improved proteins. They also find that these misfolded states have long lifespans, suggesting that they are a key component of the observed elastic, extended folding speed.

“Due to the nature of this type of misfolding, proteins get stuck,” Jiang said. “Proteins need to go back to the folding process to correct mistakes. This is time-consuming and energetically expensive. The demonstration of this mechanism helps us understand how proteins are folded and helps to give examples of how they don’t work.

In addition to Jiang, O’Brien, and Fried, the research team includes Pennsylvania graduate students Ian Sitarik and Assistant Professor of Statistics and co-authors at Johns Hopkins University, Yingzi Xia and Piyoosh Sharma.