Cellular Function and Regulation of Protein Kinases
Creators: name: University Of California Office Of The President
Publisher: name: University of California Office of the President
publisherIdentifier: https://ror.org/00dmfq477
publisherIdentifierScheme: ROR
Year: 2024

description: Abstract: Protein phosphorylation is one of the most important regulatory mechanisms in biology, and understanding how protein kinases work at atomic level resolution is one of the most topical problems in signal transduction. The goals of this project are not only to understand the molecular basis for protein kinase structure and function but also to understand how a single phosphate can regulate the structure and function of a protein kinase. To achieve this goal, we will combine the extensive protein kinase signaling expertise at UCSD with the unique neutron diffraction resources at Los Alamos. While Xray crystallography provides atomic level resolution data that is essential for the computational analysis of protein structure and function, the complete chemical profile of a kinase cannot but comprehended without having high resolution data that also defines the protons. The protons are an essential part of the networks that are mediated by protein phosphorylation. Neutron diffraction will also allow us to understand how conserved water molecules contribute directly to structure, to catalysis, to regulation, and to allostery. This information is not now available from Xray data but will be forthcoming from this novel merging of UC and LANL expertise. The experimental signaling expertise at UCSD will focus on two protein kinases, cAMP-dependent protein kinase (PKA) and the splicing kinase, SRPK. PKA, which is regulated by an essential phosphorylation of its Activation Loop, provides us with the opportunity to understand the chemical details of how a single phosphate can nucleate the entire enzyme and poise it for catalysis. It serves as a prototype for much of the protein kianse superfamily. In contrast, SRPK serves as a prototype for understanding how the structure of a phosphorylation-independent kinase is regulated. Through this unique collaboration, we will apply, for the first time, neutron crystallography techniques that visualize the role of water and phosphate in PKA and SRPK. We hypothesis that these small molecules and atoms hold key to understanding protein kinase function. We will combine the unique capabilities of LANL in neutron crystallography with the broad expertise on PKA and SRPK signaling at UCSD to solve this problem. Computational expertise, coupled with the new experimental data, will be provided by J.A. McCammon at UCSD, M. Jacobson at UCSF, and M. Wall at LANL. We believe that these highly interdisciplinary studies will pioneer new dimensions in our understanding of protein kinase function and in protein phosphorylation in general. Our proposal is an interdisciplinary collaboration between four investigators at the University of California, San Diego (Susan Taylor, Gourishankar Ghosh, J. Andrew McCammon, and Joseph Adams), Matthew Jacobson at the University of California, San Francisco, and Paul Langan and Mike Wall at the Los Alamos National Laboratory). We will train our students to use the internationally unique Macromolecular Neutron Crystallography capability at Los Alamos Neutron Science Center, which includes the Protein Crystallography Station, Biological Deuteration Laboratory, robotic expression and crystallization facilities and Computational Tools for Neutron Crystallography consortium, to determine the neutron structures of two protein kinases, PKA and SRPK, as well as on PKA regulatory proteins. The collaboration will depend heavily on our UCSD expertise and capabilities in PKA and SRPK molecular biology. Protein kinase defects are associated with many diseases and are important drug targets. Phosphorylation and ordered water molecules are central for all protein kinases but no one has yet collected neutron diffraction data for this important enzyme family. Understanding how these enzymes work and how their activity is regulated by phosphorylation has great disease relevance and will aid in rational drug design to improve therapeutics. • To build a conduit between LANL and UC campuses with graduate students and undergraduates serving as the glue. • To utilize our combined expertise to train graduate students, undergraduates, and postdoctoral fellows in the next-generation research and technology. • To build a capability for determining enzyme mechanism, MD simulation and rational drug design, by combining unique and complementary aspects of LANL and UC, that will have relevance in energy security, biothreat reduction, therapeutics, and fundamental science.