An international team of researchers has discovered how a GTP sensor kinase develops from a tumor-promoting kinase.
Led by researchers from the University of Cincinnati and Japan’s University of Tokyo, the High Energy Accelerator Research Organization (KEK), Keio University, Rikkyo University, Hoshi University, Tokai University and Kansai Medical University, the results were published on May 2 published in the magazine structure.
Atsuo Sasaki, Ph.D., one of the research team’s principal investigators, said the results provide a crucial insight that connects the dots between kinases’ ATP preference, G-protein GTP recognition and the evolutionary mechanism of nucleotide specificity and could lead to the development of a new anticancer drug targeting the GTP sensor kinase PI5P4Kβ, utilizing the discovered structural information that allows GTP to be used in place of ATP.
“Basically, kinases use ATP to phosphorylate and control their substrates, and many diseases, including cancer, are caused by the dysregulation of kinases. The ability to modulate kinase activity is central to the fight against cancer and multiple diseases,” said Sasaki, associate professor at the University of Cincinnati College of Medicine.
Kinases are a type of enzyme essential for various cellular processes including signaling, transcription and metabolism. Protein kinases, which represent the largest superfamily of more than 500 genes in the human genome, phosphoinositide kinases (PI kinases) and inositol phosphate kinases (IP kinases, including inositol kinases) share structural motifs dedicated to ATP recognition followed by hydrolysis and phosphotransfer to their substrates.
Although there is extraordinary diversity in their structure, substrate specificity, and signaling pathways involved, all of these kinases use ATP as the physiological phosphate donor. However, how and why kinases possess the ATP preference among other nucleoside triphosphates remains largely unknown, Sasaki said.
“Hence, we were surprised when we found the strong GTP preference of phosphatidylinositol-5-phosphate-4-kinase-β (PI5P4Kβ),” said Sasaki. “Our previous studies show that PI5P4Kβ acts as an intracellular GTP sensor and regulates tumorigenesis and stress responses. But we didn’t know how this particular GTP reactivity is acquired that my team set out to explore.
“To understand the mechanism of GTP dependency, it is important to understand how kinases recognize ATP. We were surprised that there was no comprehensive study of the ATP recognition mechanism by kinases,” adds Sasaki.
Sasaki said there are more than 600 kinases, including protein and phosphoinositide kinases, whose structures are solved in the ATP-bound form. However, this data is stored individually. Despite extensive efforts to target kinases for cancer therapy, the exact mechanism by which kinases recognize ATP has been unclear.
The multidisciplinary, international team was organized and first performed a structural comparison of 661 kinases and 128 G proteins that utilize GTP, revealing a common mechanism for ATP recognition by kinases. Then the research team investigated the GTP recognition mechanism of PI5P4Kβ through a combination of biochemical and structural analyzes using a nuclear magnetic resonance (NMR) activity assay, followed by an X-ray structure study.
In addition, a cutting-edge technology called Fragment Molecular Orbital (FMO) calculation enabled the team to identify the critical amino acid residues and protein-nucleotide interactions that make PI5P4Kβ a GTP-reactive kinase.
The evolutionary retrograde mutations, reversing the evolution of the GTP-using PI5P4Kβ from the ATP-using kinase PI4P5K, revealed two critical mutations in a short stretch of sequence that the researchers called the guanine efficient association (GEA) motif that endowed PI5P4Kβ with of GTP sensor activity.
“Through our multinational interdisciplinary collaboration and the team’s more than five years of hard work, we were able to understand how the dogmatic rule of ATP utilization by kinases can be overturned to GTP preference in PI5P4Kβ,” said Sasaki. “We are excited to continue our research on PI5P4Kβ to develop cancer elimination therapies that increase dependence on this GTP sensor kinase.”
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Koh Takeuchi, The GTP Responsiveness of PI5P4K? is developed through a tradeoff between activity and specificity., structure (2022). DOI: 10.1016/j.str.2022.04.004. www.cell.com/structure/fulltex … 0969-2126(22)00131-9
Provided by the University of Cincinnati
Citation: Research team unravels the trick of evolving the GTP sensor (2022, May 2), retrieved May 2, 2022 from https://phys.org/news/2022-05-team-unravels-evolving-gtp-sensor. html
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