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Molecular Motors Similar To Macro Engines

Think of individual proteins as strings of spaghetti. When they fold and combine with other proteins, they create tools that do important work inside a body. These tools are known as molecular machines.

Protein folding is a “carefully orchestrated sequence of events,” says Lars Konermann, a professor in Western University’s Department of Chemistry. “If this sequence gets interrupted, bad things can happen.” “Bad things” include a variety of illnesses, which is why Konermann and his colleagues examine the phenomenon in their labs at Western.

All types of protein motion interest Konermann. “They wiggle and they shake and fold and unfold all the time,” he explains. “Many of these movements are related to function.”

Protein movements happen in fractions of a second. Scientists can acquire “still photos” using interrogation techniques like X-ray crystallography, but “that’s not the biologically active form of the protein,” Konermann says. “We have to devise clever strategies to see what a protein does when working in its natural environment.”

Enter hydrogen/deuterium exchange mass spectrometry (HDX/MS). “Mass spectrometry is a fancy way for saying we weigh molecules. We can extract detailed information from these mass measurements.”

Konermann’s colleague, Professor Stanley Dunn in the Department of Biochemistry at Western University, is an expert on proteins involved in cellular energy conversion. Together, the Dunn and Konermann research teams recently embarked on the first in situ investigation of a catalytically active rotational molecular motor using data from both X-ray crystallography and HDX/MS. They developed techniques that allowed them to interrogate a well-known molecular motor known as ATP synthase while it worked in native biological membranes.

ATP synthase forms ATP molecules, the “energy currency of the body.” When the ATP molecule breaks apart, it releases energy that humans use for any imaginable activity, from cell formation to hair growth to raising a finger and beyond. Every day, the human body produces and consumes many kilograms of ATP.

Before proceeding with this interrogation, “we had some idea of what we might find, but the largest effect we saw was quite unexpected and novel. We had to develop new theories to make sense of our observations.”

Their interrogations of ATP synthase revealed that this nanoscale motor has lots in common with much larger manmade motors.

For instance, an ATP synthase protein takes the form of an asymmetrical shaft (like a car’s crankshaft), part of which spins inside a plain journal bearing. As it turns, the shaft’s rough surface (picture the surface of a cauliflower) grinds against its bearing. Konermann and Dunn presumed the spinning protein faced resistance.

The researchers used this crankshaft analogy to form their next question: “As this thing rotates and grinds against friction, we wanted to see where and when parts of this protein undergo deformation.”

HDX/MS data acquired during rotation revealed the presence of stress on the shaft, but this stress did not manifest itself where it was expected. As the protein grinds against the bearing segment, an “interlocking” phenomenon holds back the rotation of the protein, deforming it where the friction occurs.

“The stress in this part accumulates. It stays under stress all the time. It’s like a rubber band gets twisted, then lets go a little, then gets twisted a bit more, so that it always maintains its twist.” These findings were reported in a recent issue of the journal Proceedings of the National Academy of Sciences of the United States of America.

These surprises occurred while interrogating a well-understood protein that the body can’t do without. Other researchers will be able to use this in situ HDX/MS interrogation technique when they investigate proteins that aren’t as well understood as ATP synthase, including those that form illness-related molecular machines.

When Konermann explains the importance of his work, he talks about the value of flashlights to cave explorers. To scientists who research diseases like Parkinson’s, Alzheimer’s and cancer, every unexplored protein is a cave where they can use the novel techniques as flashlights.

“What we do in the lab is pretty fundamental. It is essential to understand these basics before you can tackle high-level questions related to disease and drug action mechanisms,” Konermann says. Dunn notes that “other scientists have decried the shortage of structural techniques that can be applied to proteins in their natural environments. Our work provides a novel method for these studies and our surprising results demonstrate the value of pursuing this type of approach.”

Professor Lars Konermann leads the exploration of the folding mechanisms of proteins, their conformational dynamics, and their interactions with other molecules at Western University’s Departments of Chemistry and Biochemistry. He and his group are developing and applying novel electrospray mass spectrometry (ESI-MS) techniques for this purpose. This research bridges the areas of biophysics, biochemistry, and analytical chemistry. Distinguished University Professor Stanley Dunn’s work is focused on protein structure and function, with emphasis of the structure and mechanism of ATP synthase. Other areas of current interest include the upregulation of energy production in support of T cell activation in the immune response and the analysis of protein-protein interactions in the development of amyotrophic lateral sclerosis (ALS).

This article originally published on the Western Science website.

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