NMR reveals entropy redistribution: Bacterial proteins

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  • Published: May 1, 2017
  • Author: David Bradley
  • Channels: NMR Knowledge Base
thumbnail image: NMR reveals entropy redistribution: Bacterial proteins

Form over function

Credit: CDC The scientists conducted their experiments in Staphylococcus aureus, a common cause of skin, sinus and lung infections.

Research carried out at the of the METACyt Biomolecular NMR Laboratory at Indiana University Bloomington has revealed evidence that tiny changes in atomic movements in bacterial proteins can play a big role in how these microorganisms function and evolve.

According to senior author David Giedroc and his colleagues, allosteric communication between two ligand-binding sites in a protein is critical to biological regulation but exactly how it works is unclear. The new work, published recently in the journal Proceedings of the National Academy of Sciences takes science down a different route to home in on an explanation. The findings counter the conventional perspective that a protein's shape, or tertiary and quaternary structure are the most important factor in deriving function from form and suggest that there is another dimension at play in evolution and in protein behaviour within organisms.

"This study gives us a significant answer to the following question: How do different organisms evolve different functions with proteins whose structures all look essentially the same?" explains Giedroc, who is Lilly Chemistry Alumni Professor in the IU Bloomington College of Arts and Sciences' Department of Chemistry. "We've found evidence that atomic motions in proteins play a major role in impacting their function." Essentially, entropy is important to protein form and function.

Entropic insight

"What we've shown is atomic-level motional disorder - or entropy - can impact gene transcription to affect the function of proteins in major ways, and that these motions can be 'tuned' evolutionarily," adds post-doctoral researcher Daiana Capdevila. "This may allow bacteria to rapidly evolve new ways to overcome medical treatment since atomic motions can be optimized for function more easily than a physical structure."

The ongoing battle between bacterial infection and modern medicine one of the important strategies constantly undergoing development is to "map" enemy territory. If biomedical science can extract the molecular structure of a protein involved in blocking immune system activity, for instance, it might be possible to develop new pharmaceuticals that stave off drug resistance. However, mapping protein territory often assumes that a protein's shape is fundamental to its function, and this new work suggests that this axiom if not completely fake news is at least in some ways an economy of the complete picture. In one particular sense, the concept of protein function and form often makes the simplifying approximation that a protein is essentially rigid, a point reinforced perhaps by X-ray crystallographic structural studies.

The new study provides an insight into this new perspective that allows microorganisms, such as Staphylococcus aureus a cause of skin, sinus and other more serious infections, especially in its multiple-drug-resistant form, MRSA, respond to the body's efforts, through the immune response to fight the infection. MRSA, tuberculosis, Clostridium difficile, E coli O157 and other bacteria with significant resistant strains spreading widely represent a serious healthcare risk in the face of poorly performing antibiotics and a lack of novel drugs in this area emerging from pharmaceutical R&D pipelines. For instance, almost half a billion people worldwide develop multidrug-resistant tuberculosis each year, for example, according to the Centers for Disease Control and Prevention. This and many other bacterial infections represent an enormous problem for healthcare to address.

Zinc spike

Giedroc and colleagues analysed the protein CzrA (chromosomal zinc-regulated repressor) using nuclear magnetic resonance (NMR) spectroscopy. This protein controls how bacteria regulate zinc levels and so resist the human immune system. If the immune system floods the site of infection with zinc ions that can limit bacterial growth, equally starving the bacteria of this essential element can kill them too. CzrA controls the bacterial "zinc pump" which pushes zinc out of the bacterium if too much is present because of the immune system's efforts to poison the microbes with metal.

The NMR spectra reveal hotspots in the CzrA where sensitive atoms reside, swapping out the atoms by changing amino acids in these hotspots and so altering the entropy of the protein. The changed amino acids are not necessarily in the active site of the protein and so off the "map", can dramatically alter its ability to regulate zinc.

"There's no way anyone could have predicted these areas played a role in zinc regulation by simply looking at the protein structure," Giedroc explains. "Once you know where these 'hotspots' are located, however, it's theoretically possible to design a small molecule or drug to produce the same effect as our amino acid-swapping experiment - that is, to essentially shut off the protein." Such "allosteric drugs," affect the hotspots in a protein without directly targeting the protein's active site. This new approach goes beyond form and function and exploits the role of atomic motion in function with a long-term goal to develop new approaches to interfering with proteins in bacterial infection.

"The next step in the project is to understand the degree to which our entropy redistribution model underpins allostery in other transition metal-sensing proteins that look like CzrA yet impact adaptation by bacterial pathogens to other host-derived stresses," Giedroc told SpectroscopyNOW.

Related Links

Proc Natl Acad Sci 2017, 114, 4424-4429: "Entropy redistribution controls allostery in a metalloregulatory protein"

Article by David Bradley

The views represented in this article are solely those of the author and do not necessarily represent those of John Wiley and Sons, Ltd.

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