"Many essential cellular processes are carried out by "macromolecular machines", large assemblies that often include tens of different proteins, each making a specific contribution to the function of the machine. Characterization of these large macromolecular complexes using a combination of techniques constitutes the next frontier in structural biology. Interestingly, despite including many different proteins, macromolecular complexes often display minimal enzymatic activity. This suggests that they truly function as macromolecular machines, in which function is related to "mechanical" changes that affect interactions and activity. Characterizing different states and conformations of a macromolecule, and correlating these findings to biochemical and functional information, can be essential to establish the mechanism of these remarkable cellular machines."
The long-term goal of our research is to determine the molecular principles and logic that underlie transcriptional regulation in humans. As a model system, we studying the human steroid receptors, how they assemble at complex promoter sequences, and the relationship between these interactions and cellular outcomes. Our group uses thermodynamic approaches to experimentally dissect receptor- promoter binding energetics and statistical thermodynamics to synthesize overall behavior; we collaborate closely with Dr. James Lambert¹s lab (UCDenver Pathology) to link these findings to in vivo behavior.
Ion channels are basic molecular elements responsible for the generation of cellular electricity. The opening and closing (gating) of these transmembrane proteins give rise to ionic fluxes that generate electrical signals in tissues all across the body. The generation of these signals is critical for phenomena as diverse as nerve action potentials, sensory transduction, pain sensing, muscle contraction, hearing, vision, and hormone secretion. Ion channels gate in response to external stimuli such as binding of ligands (ligand-gated ion channels) or changes in transmembrane electric field (voltage-gate ion channels). Our research is focused on understanding fundamental molecular mechanisms of ion channel function. This includes studying the basic structural elements involved in coupling a stimulus to a change in state (from close to open for instance) as well as studying how ion channels function as larger macromolecular complexes. To attack these questions, we use a combination of electrophysiology, biochemistry, fluorescence imaging, electron paramagnetic resonance spectroscopy, and structural biology. Trainees in the lab would be encouraged to take projects that would provide expertise in more than one of these approaches.
Structural analysis of protein glycosylation
using NMR and mass spectrometry.
Research in the Catalano lab focuses on molecular mechanisms of virus assembly in the double-stranded DNA viruses. We couple detailed enzyme kinetic analyses with biophysical and structural interrogation of the molecular motor that "packages" the viral genome into a capsid shell. Maturation of the nucleoprotein particle into an infectious virus is examined in defined reaction mixtures. The lambda capsid system is further harnessed for the construction of "designer" nanoparticles for use as therapeutic and diagnostic agents.
We are committed to advancing individual medicine by examining the unique biology of an individual to assess truly personalized treatments. Our state of the art facility provides integrated solutions to systems biology and is located at Colorado's Fitzsimons Bioscience Park.
My lab is interested in understanding the molecular basis of essential processes that regulate gene expression. We use biophysical, biochemical methods, and structural methods, including X-ray crystallography. Our insights into these fundamental mechanisms will contribute to a better understanding and ability to regulate gene expression processes involved in human diseases from cancer and heart disease to bacterial infections and will assist in drug development efforts.
Omics technologies, especially metabolomics and proteomics, have helped us revealing emerging patterns in systemic responses to acute or chronic hypoxia. By focusing on cancer metabolism and (red) blood cell biology, we are increasingly appreciating shared molecular mechanisms driving systemic responses to trauma/ hemorrhagic shock, I/R injury, sickle cell disease, ageing and inflammation, mammalian hibernation and pulmonary hypertension.
Metal ions are essential nutrients in all forms of life. Despite their important roles, metals can be toxic and elicit different kinds of immune responses, causing diseases. Chronic beryllium disease is a fibrotic lung disorder caused by beryllium exposure, while nickel ion is the dominant allergen for contact dermatitis.
Our major goal is to understand the mechanisms of the metal containing ligands for alpha/beta TCRs from metal reactive human T cells. Using structural biology we will be able to discern the respective rule for metal binding and find potent compounds to inhibit metal binding, thus abrogating T cell responses. We are also interested in the molecular basis of metal induced autoimmunity.
Recently we observed metal ions directly attenuate the binding affinity between TCR and pMHC interaction. Our goal is to understand how the metal ions attenuate the TCR/pMHC binding affinity and what effects they have on T cells activation. The results can help to elucidate the molecular basis for some autoimmunity caused by metals. We are interested in redox signaling in chloroplast and T Cells.
We are studying a unique iron-sulfur enzyme, ferredoxin: thioredoxin reductase, containing both a catalytic [4Fe-4S] cluster and a redox active disulfide. The overall objective of this proposal is to decipher the mechanisms of reduction of disulfide catalyzed by iron sulfur cluster and thiol-disulfide exchange reactions in ferredoxin /thioredoxin system. Redox signaling in apoptosis involves the oxidation and reduction of cysteine residues in critical thiol proteins. Changes in the structure and activity of these proteins can initiate cell responses, or modify the response of cells to other signals.
My proposed research focuses on the regulatory mechanisms of apoptosis signal-regulating kinase 1 (ASK1) mediated T cell apoptosis. ASK1 is a member of the mitogen-activated protein kinase kinase kinase (MAP3K) family, which activates both the SEK1-JNK and MKK3/6-p38 MAP signaling cascades and constitutes a pivotal signaling pathway in cytokine- and stress-induced apoptosis. This signaling pathway mediates a variety of cellular events, including cell proliferation, differentiation, and death.
We are interested in understanding multiple molecular interactions that go awry during both inflammatory diseases and cancer progression. The novelty in our group’s approach is that we utilize highly integrative methods to probe interactions from atomic resolution techniques to cell-based techniques. When cellular and clinical studies are combined with molecular and biochemical studies, a complete understanding of the particular system under study can be drawn.
We are focused on developing and utilizing strategies for the detection and characterization of proteins in health and disease. Our goal is to understand underlying mechanisms of disease at the molecular level using mass spectrometry as our primary analytical tool.
Using cell culture models we are performing studies to identify ECM components and modifications that influence metastatic potential of breast epithelial cells. We are also developing quantification methods to identify mediators of multiple organ failure in shock models and patients.
Synthetic peptide and antipeptide approaches play major roles in understanding protein structure and function.
The basic and translational research focus of my laboratory is on two areas. The first is the roles of complement receptors and membrane regulatory proteins in the immune response, with a special emphasis on B lymphocytes and autoimmune diseases. The second is the role of autoantibodies and the evolution of autoimmunity in RA from the pre-symptomatic autoantibody-positive period through the onset of clinically active disease.
Development and application of advanced computational techniques for biomedicine, particularly the application of statistical and knowledge-based techniques to the analysis of high-throughput data and of biomedical texts. Also, neurobiologically and evolutionarily informed computational models of cognition, and ethical issues related to computational bioscience.
My laboratory is currently focused on knowledge-driven extraction of information from the primary biomedical literature, the semantic integration of knowledge resources in molecular biology, and the use of knowledge in the analysis of high-throughput data.
Our work focuses on the formation and regulation of chromatin domains and their ultimate roles in the nucleus. We are particularly interested in the mechanisms of heterochromatin establishment and function. Heterochromatin operates in organisms from yeast to humans to determine cell identity and maintain genome stability by silencing genes. Because heterochromatin functions in such central processes, misregulation of this genomic structure can have dire consequences such as cancer or abnormal development. Our work investigates the mechanisms by which silencing is carried out. We use a combination of in vitro assembly of chromatin domains, mechanistic biochemistry, proteomic analysis, and genome-wide chromatin profiling to understand the complex superstructural “neighborhoods” of chromosomes.
The action of small molecules at receptors and other proteins in signalling cascades leads to major changes in behavior. These small molecules act by producing a change in protein structure and dynamics that ultimate leads to changes in nueronal signalling. Our research focuses on two different classes of modulators of neuronal signal transduction, namely alcohols and pheromones. Alcohols act on a variety of receptors and other neuronal proteins, and lead to pharmacological changes that can result in alcohol intoxication and alcohol dependency.
Atopic Dermatitis, Autoimmunity/Rheumatology, Basic Immunology, Cancer, Chronic Beryllium Disease, Genetics, Immunobiology, Molecular Immunology, Type-1 Diabetes, Structural Biology
Our research focuses on understanding the structure and function of the most versatile of all biological macromolecules: RNA. Our main focus is to understand RNAs produced and used by viruses to take over an infected cell. We are motivated by two premises: (1) infectious diseases (such as viruses) are still the largest threat to human health worldwide, and (2) by studying how viruses take over the cell’s machinery, we learn a tremendous amount about fundamental processes within the cell itself.
Research in my group focuses on the molecular mechanisms of epigenetic regulation and phosphoinositide signaling. We apply high field NMR spectroscopy, X-ray crystallography and a wide array of biochemical and molecular biology approaches to characterize the atomic-resolution structures and functions of chromatin- and lipid-binding proteins implicated in cancer and other human diseases.
The LaBarbera laboratory is focused on drug discovery and development targeting cancer and diabetes. To accomplish this we utilize a multidisciplinary approach encompassing assay development for high-throughput screening (HTS) and confocal image based high-content screening (HCS), natural products small molecule library development, mechanism of action studies, and drug design and medicinal chemistry. The LaBarbera lab has pioneered techniques in validating and implementing 3D-tissue culture models of human disease for HCS/HTS,including: human lens epithelial spheroids (lentoids) for diabetic eye disease research and the multicellular tumor spheroid (MCTS) model for cancer research. We couple these models with surrogate biomarker reporters for phenotypic screening to identify small molecule bioactive modulators of human disease. Once we identify lead compounds we determine and validate their molecular target(s) and characterizethe mechanism(s) of action using in silico, in vitro, cell based, and in vivo models to design more potent “druglike” lead compounds with a long-term goal of clinical translation.
Our lab is interested in developing mass spectrometry and computational methods to understand protein dynamics in health and disease. Current projects include developing multi-omics strategies to quantify protein alternative isoforms; investigating the role of ER stress and protein glycosylation in cellular stress responses; and identifying the trends and focuses of research topics in the biomedical literature.
Muscular dystrophy (MD) refers to a group of degenerative muscle diseases that cause progressive muscle weakness. MD affects all types of muscles. For example, decreased function of heart muscles causes heart diseases that include cardiomyopathy and congestive heart failure. At present there is no cure available for MD, although certain palliative treatments are available to ease the pain associated with MD. Duchenne MD (DMD) and Becker MD (BMD) are two prominent types of MD, which are caused by the deficiency of a vital muscle protein known as dystrophin.
Our research focuses on identifying molecular mechanisms underlying the assembly of macromolecular complexes, with a focus on multisubunit complexes formed by septin proteins. All cellular processes require the function of multisubunit complexes, and while much attention has been given to solving the final structures of such assemblies, comparatively little is known about how individual subunits adopt oligomerization-competent conformations and find their partner subunits in the crowded, dynamic cellular milieu. Our favorite multisubunit assembly is the septin protein complex, found in nearly all eukaryotes and required for a variety of cellular processes. Our studies challenge the idea that thermodynamics (i.e., the affinities between proteins randomly colliding in solution) is the main driving force of protein complex assembly, and favor largely unappreciated roles for the kinetics of protein folding and cooperativity between protein-protein interactions.
The Musselman laboratory is interested in the structural basis of chromatin signaling. Chromatin is a complex between genomic DNA and histone proteins. The basic subunit of chromatin is the nucleosome, which consists of an octamer of histone proteins wrapped by ~147 base pairs of DNA. Protruding from the wrapped octamer are intrinsically disordered N- and C-terminal regions of the histones, referred to as the histone tails. A plethora of post-translational modifications (PTMs) has been discovered to exist on the histone tails. These PTMs are thought to act in combination to signal for structural rearrangements in chromatin that ultimately lead to genome regulation. We are interested in the molecular underpinnings of how these PTMs are “read-out” by histone binding domains (or reader domains). Though much work has been done to determine this in the context of histone peptide fragments, very little is understood about how histone PTMs function in the context of the nucleosome. We are using NMR spectroscopy to investigate histone tail conformation in the context of the nucleosome, to determine how reader domains bind nucleosomal histone tails, and to determine how various modes of modulation of the nucleosome regulates these signaling events.
Protein, RNA, and other functional molecules that exist in living organisms are the product of millions of years of evolution. The substitutions that have occurred over the years had to have been compatible with the constraints of structure and function, and thus the evolutionary record provides critical data for understanding macromolecular structure/function/sequence relationships.
Regulation of genome access underlies growth, development, differentiation, and disease states including tumor initiation. Genome access is regulated by dynamic chromatin landscapes. We want to uncover fundamental mechanisms that shape chromatin landscapes by mapping chromatin structure at high temporal and spatial resolution using new experimental and/or computational methods. This research program in chromatin biology is at the intersection of genomics, biochemistry, and structural biology.
Dr. Reisdorph's research is designed to integrate clinical proteomics, metabolomics, and bioinformatics in order to develop and customize clinically relevant methodologies to diagnose or monitor disease states. Results from these studies will also significantly enhance the knowledge of the biochemical mechanisms of diseases. In addition, Dr’s Reisdorph’s facility will specialize in post-translational modification analysis as well as the identification of differentially regulated proteins from a variety of sources including cell extracts, biofluids, and tissue samples. Dr. Reisdorph uses a variety of techniques in her work, including two-dimensional gel electrophoresis, DiGE, tandem mass spectrometry, and quantitative labeling and non-labeling strategies. Dr. Reisdorph organizes proteomics hands-on workshops and web-based courses through National Jewish and the University of Colorado Denver. Since 2005, Dr. Reisdorph and her team have instructed almost a dozen 3-4 day workshops, for a total of over 120 individuals, who come from a variety of backgrounds. Dr. Reisdorph is currently expanding her training program to include additional hands on courses, such as quantitative proteomics, and distance-learning courses, such as database searching.
We are interested in understanding the molecular mechanisms and physiological functions of transporters and ion channels. Our current research focuses on the mitochondrial calcium uniporter and the mitochondrial NCLX protein, which mediate calcium uptake and efflux in mitochondria, respectively. These transport proteins regulate key cellular processes, such as ATP production and cell death, and their malfunction has been linked to neurodegenerative disease and death of cardiomyocytes following myocardial infarction. Our research is problem-driven — trainees will work with the PI to identify key questions, and address these using a wide range of tools available in the lab, including membrane-protein biochemistry/reconstitution, electrophysiology, structural biology, genome editing, and imaging.
Research in the Tucker Lab focuses on development of technologies for manipulation and probing of protein interactions and pathways. In particular, we are focusing on the emerging area of cellular optogenetics, developing pioneering new engineered protein tools to precisely regulate cell function with light.
Protein and nucleic acid architecture is defined by the 3D average structure, but biomolecules are inherently dynamic systems. Using NMR, we contribute to the understanding of communication networks by modeling realistic ensembles of structures, identification of allosteric mechanisms and interactions with other proteins, nucleic acids or small ligands. An important pillar of this program is the establishment of our recently developed exact nuclear Overhauser enhancement (eNOE) methodology for a realistic representation of molecular spatial sampling.
We aim at atomic-scale descriptions of the following mechanisms: i) Motor adaptors in complex dynamic protein assemblies associated with the motor cytoplasmic dynein: We conduct experiments with the C-terminal tail of the dynein light intermediate chain to provide molecular insight into how the elongated, flexible scaffold engages with functionally diverse dynein adapters. ii) Structural landscape of human peptidyl-prolyl cis/trans isomerase Pin1 allostery: How does the WW domain transmit information from its binding site to the spatially separated catalytic PPI site? iii) Long-range coupling networks in PDZ domains: An NMR-based evidence of the presence of correlated motion along the proposed pathways would constitute a major advance both in NMR methodology, as well as towards an understanding of dynamics-function relationships and allostery. iv) Allostery in protein-RNA interaction: The elucidation of the allosteric mechanism of RsmE upon binding of non-coding RNA RsmZ may uncover more general principles on how RNA binding domains in tandem or homodimeric RNA binding proteins fine tune their RNA binding affinity.
My laboratory has two major research focuses. The first is to understand the molecular mechanism of pre-mRNA splicing using a combination of X-ray crystallography, molecular biology, biochemistry and yeast genetics approaches. Splicing of pre-mRNA is essential for gene expression in all eukaryotes. In higher eukaryotes such as mammals, an average of 95% of the nucleotides in the primary transcript (pre-mRNA) of a protein-encoding gene are introns.
We are interested in the broad field of structure and function relationship of membrane protiens and membrane protein complexes. Right now, our focus is on the mechanism of protein translocation between cytosol and mitochondria. We use primarily X-ray crystallography and Cry-Eletron Microscopy to explore the structural characteristics of the corresponding membrane proteins, so to understand their biological roles in the cell.