Kevin Dalby's Lab

Featured research (6)

The luminescence method measures the spontaneous emission of a photon by a substance. Among the various types of luminescence, fluorescence and chemiluminescence are the most dominant methods used in high throughput screening (HTS) campaigns due to their keen sensitivity, broad linearity, robustness, and flexible automation/miniaturization. Fluorescence and chemiluminescence result from excited state photons, but they differ in how the photons are excited. In fluorescence, the energy to excite photons is provided by an external light source, which offers enhanced sensitivity and the ability to tune the excitation wavelength for diverse targets of interest. Because of the need for an external source of light, high background, and significant interference with chemical libraries can be problematic in fluorescence-based HTS. In chemiluminescence, the energy is provided by a chemical reaction, often involving oxygen. Chemiluminescence does not require radiation and has become an indispensable detection method. One particular form of chemiluminescence is called bioluminescence, where an enzyme catalyzes the chemical reaction, most often a luciferase enzyme. Early, bioluminescence-based technology measured transcription, cell viability, or ATP-dependent biochemical processes. This chapter addresses new and ongoing developments, such as the utilization of new luciferases, and the caging of luciferins, and their application in high throughput screens. It also addresses the use of proximity-based technologies such as BiLC (Bimolecular luciferase complementation), BRET (Bioluminescence Resonance Energy Transfer), and Alpha (Amplified Luminescence Proximity Homogenous Assay) to quantify changes in analyte concentration, biomolecular interactions, and post-translational modifications in such screens.
Translation is a highly energy consumptive process tightly regulated for optimal protein quality and adaptation to energy and nutrient availability. A key facilitator of this process is the α-kinase eEF-2K that specifically phosphorylates the GTP-dependent translocase eEF-2, thereby reducing its affinity for the ribosome and suppressing the elongation phase of protein synthesis. eEF-2K activation requires calmodulin binding and auto-phosphorylation at the primary stimulatory site, T348. Biochemical studies have predicted that calmodulin activates eEF-2K through a unique allosteric process mechanistically distinct from other calmodulin-dependent kinases. Here we resolve the atomic details of this mechanism through a 2.3 Å crystal structure of the heterodimeric complex of calmodulin with the functional core of eEF-2K (eEF-2K TR ). This structure, which represents the activated T348-phosphorylated state of eEF-2K TR , highlights how through an intimate association with the calmodulin C-lobe, the kinase creates a spine that extends from its N-terminal calmodulin-targeting motif through a conserved regulatory element to its active site. Modification of key spine residues has deleterious functional consequences.
The SARS-CoV-2 spike protein is a critical component of vaccines and a target for neutralizing monoclonal antibodies (nAbs). Spike is also undergoing immunogenic selection with variants that increase infectivity and partially escape convalescent plasma. Here, we describe Spike Display, a high-throughput platform to rapidly characterize glycosylated spike ectodomains across multiple coronavirus-family proteins. We assayed ∼200 variant SARS-CoV-2 spikes for their expression, ACE2 binding, and recognition by 13 nAbs. An alanine scan of all five N-terminal domain (NTD) loops highlights a public epitope in the N1, N3, and N5 loops recognized by most NTD-binding nAbs. NTD mutations in variants of concern B.1.1.7 (alpha), B.1.351 (beta), B.1.1.28 (gamma), B.1.427/B.1.429 (epsilon), and B.1.617.2 (delta) impact spike expression and escape most NTD-targeting nAbs. Finally, B.1.351 and B.1.1.28 completely escape a potent ACE2 mimic. We anticipate that Spike Display will accelerate antigen design, deep scanning mutagenesis, and antibody epitope mapping for SARS-CoV-2 and other emerging viral threats.
Extracellular signal–regulated kinase (ERK) is a mitogen-activated protein kinase (MAPK) that mediates cellular processes such as proliferation, differentiation, cell motility, and survival. Dysregulation of the ERK signaling pathway is believed to have a protumorigenic role in many cancers, and studies also implicate it in a variety of other proliferative diseases. Within the ERK signaling pathway, protein-protein interactions via enzyme-docking sites help generate signal specificity and direct ERK to subsequent binding partners or substrates. ERK possesses two known docking sites that are distinct from its catalytic site: the D- and F- recruitment sites (DRS and FRS). Over time, our group has characterized these sites through a combination of structural and kinetic studies, including computational and biochemical techniques, centering around a model ERK substrate EtsΔ138 (residues 1–138 of the transcription factor Ets-1). These studies are part of a growing effort to elucidate new insights into ERK signaling and to evaluate the role of each binding site in specific ERK interactions. Furthermore, the development of inhibitors that target these docking sites offers a way to impede both catalytic and noncatalytic functions of ERK, which may provide therapeutic benefit in disease states driven by ERK signaling. Here, we describe the features of the DRS and FRS of ERK, their roles in the phosphorylation of EtsΔ138, and the status, mechanisms, and implications of targeting these sites with inhibitors.

Lab head

Kevin Dalby
  • Division of Chemical Biology & Medicinal Chemistry
About Kevin Dalby
  • Dr. Kevin Dalby UT-Austin (Twitter @KevinNDalby) is a professor of chemical biology and medicinal chemistry in the College of Pharmacy, and Department of Oncology at The University of Texas in Austin. He studies mechanisms of cell signaling to develop targeted therapeutics. Dr. Dalby's research areas include biochemistry, cancer, cell biology, chemical biology, drug discovery & diagnostics, and enzymology.

Members (4)

Eun jeong Cho
  • University of Texas at Austin
Juhoon Lee
  • University of Texas at Austin
Rachel Sammons
  • University of Texas at Austin
Eric A Kumar
  • Rice University
Kimberly Long
Kimberly Long
  • Not confirmed yet