A5048362510- Superconducting Materials and Applications
- Enzyme Structure and Function
- Protein Structure and Dynamics
- Ubiquitin and proteasome pathways
- Crystallization and Solubility Studies
- X-ray Diffraction in Crystallography
- Metalloenzymes and iron-sulfur proteins
- Electron and X-Ray Spectroscopy Techniques
- 14-3-3 protein interactions
- Advanced Electron Microscopy Techniques and Applications
- Peptidase Inhibition and Analysis
- Genetic and Kidney Cyst Diseases
- Microbial Natural Products and Biosynthesis
- Cellular Mechanics and Interactions
- Cardiomyopathy and Myosin Studies
- Antimicrobial Peptides and Activities
- Counseling Practices and Supervision
- Advanced Proteomics Techniques and Applications
- Hedgehog Signaling Pathway Studies
- Biochemical and Molecular Research
- Heat shock proteins research
- RNA and protein synthesis mechanisms
- Genetic Syndromes and Imprinting
- Enzyme Catalysis and Immobilization
- Folate and B Vitamins Research
Brigham Young University
2008-2025
Montana State University
2016-2023
Howard Hughes Medical Institute
2017
University of Washington
2011-2017
University of New Orleans
1977
Ducks Unlimited
1977
East Sussex County Council
1909
Royal Albert Edward Infirmary
1909
Trenton Psychiatric Hospital
1898
London Borough of Islington
1898
Significance We describe an approach to blocking protein–protein interactions in living cells and use it probe the earliest stages of epigenetic regulation stem cell differentiation. a computationally designed protein that tightly binds EED disrupts PRC2 function both cancer cells. Expression binder at different identifies first critical repressive H3K27me3 mark embryonic development.
Abstract TNK1 is a non-receptor tyrosine kinase with poorly understood biological function and regulation. Here, we identify dependencies in primary human cancers. We also discover MARK-mediated phosphorylation on at S502 that promotes an interaction between 14-3-3, which sequesters inhibits its activity. Conversely, the release of from 14-3-3 allows to cluster ubiquitin-rich puncta become active. Active induces growth factor-independent proliferation lymphoid cells cell culture mouse...
The covalent fusion of the sterile alpha motif domain from human translocation ETS leukemia protein (TELSAM) with a interest has been shown to significantly improve ability create crystals and enhance diffraction quality. TELSAM, acting as chaperone that forms six-subunit-per-turn helical polymer at low pH, creates stable crystal lattice when attached another protein. However, challenges persist in choosing best linkers between specifically 1TEL variant, target protein, current methods...
TELSAM refers to the sterile alpha motif domain (SAM) of human translocation ETS leukemia protein (TEL). is known spontaneously polymerize into a 6-fold helical polymer in low pH. was previously used as crystallization chaperone fused CMG2 vWA using Threonine–Valine linker. This 1TEL-TV-vWA construct crystallized and mounted house, then analyzed Synchrotron X-ray source. The structure solution this resulted consistently high R-values. Analysis recorded reciprocal lattice showed three...
Crystallization is the rate-limiting step in macromolecular X-ray diffraction studies. Usually, crystallization methods are only successful for a fraction of known proteins, producing single crystal with high resolution time-consuming, expensive, and laborious process. TELSAM protein chaperone that enhances propensity. We investigated effects target loading histidine tag inclusion on dynamics quality. tested displaying 2, 3, or 6 copies 9 kDa UBA domain 19 vWa per turn polymer. used flexible...
The [FeFe]-hydrogenase protein plays a key role in the biosynthesis of H2 various anaerobic microorganisms and green algae. This is done through sequential mechanism reactions driven by multiple radical SAM enzymes to form organometallic H-cluster found [FeFe]-hydrogenases. There currently convincing evidence supporting proposed chemical synthesis, however there still large gap knowledge concerning structural characterization [FeFe] hydrogenase complex. study aims obtain high resolution...
The TELSAM protein polymer is an efficient crystallization chaperone when fused to a variety of target proteins, but it faces some unique challenges. One these the tendency constructs form aggregates due premature polymerization unit. In past, we employed one pH-dependent trigger into TELSAM: V112E. To optimize fusion technique, introduced second mutation: L95E. We then crystallized alone and have begun crystallizing three TELSAM—TNK1.UBA connected with varying linkers. Two exhibited limited...
Many cancer cells utilize autophagy to survive chemotherapy. Understanding mechanisms in the context of therapy is important. Vacuolar protein sorting 34 (VPS34) a key regulator initiation autophagosome formation. Current studies are exploring autophagic inhibitors as potential supplements heighten responsiveness established treatments. However, many these lack specificity and have off-target effects. To date, there no FDA- approved VPS34 antagonist. RD-I-53 highly selective inhibitor VPS34....
The fusion of the sterile alpha motif domain human translocation ETS leukemia protein (TELSAM) to a interest has been shown significantly enhance crystallization propensity and improve diffraction quality. TELSAM is pH-dependent, polymer-forming chaperone which, when covalently fused interest, forms stable homogenous crystal lattice. However, despite its success, challenge persists in significant impact linkers between on quality, with best linker selection relying trial-and-error methods...
Crystallization is the rate-limiting step in macromolecular X-ray diffraction studies. Usually, crystallization methods are only successful for a fraction of known proteins, and producing single crystal with high resolution time-consuming, expensive, laborious process. TELSAM protein chaperone which enhances propensity. We investigated effects target loading histidine tag inclusion on dynamics quality. tested displaying 2, 3, or 6 copies 9 kDa UBA domain 19 vWa per turn polymer. used...
Mutations in dystrophin lead to Duchenne muscular dystrophy, which is among the most common human genetic disorders. Dystrophin nucleates assembly of dystrophin–glycoprotein complex (DGC), and a defective DGC disrupts an essential link between intracellular cytoskeleton basal lamina, leading progressive muscle wasting. In vitro studies have suggested that phosphorylation may affect interactions with actin or syntrophin, yet whether this occurs vivo affects protein function remains unknown....
While conducting pilot studies into the usefulness of fusion to TELSAM polymers as a potential protein crystallization strategy, we observed novel properties in crystals two TELSAM-target fusions, follows. (i) A can crystallize more rapidly and with greater propensity than same target alone. (ii) fusions be crystallized at low concentrations. This unprecedented observation suggests route proteins that only produced microgram amounts. (iii) The themselves need not directly contact one another...
Computational enzyme design has made great strides over the last five years. Traditional methods of require synthesis and evaluation many mutations. emerged as a powerful tool to predict how specific mutations modify protein’s activity, stability, and/or selectivity. Such computational approaches can evaluate reduce load in vitro work by identifying likely accomplish objectives. explore mutational spaces inaccessible traditional mutagenesis. cost time compared with experimental approaches....
Radical S-adenosyl-l-methionine (SAM) enzymes are ubiquitous in nature and carry out a broad variety of difficult chemical transformations initiated by hydrogen atom abstraction. Although numerous radical SAM (RS) have been structurally characterized, many prove recalcitrant to crystallization needed for atomic-level structure determination using X-ray crystallography, even those that crystallized an initial study can be recrystallize further structural work. We present here method...
Abstract Cell-cell and cell-substrate adhesions are sites of dramatic actin rearrangements where actin-membrane connections tightly regulated. Zyxin-VASP complexes localize to cell-cell adhesion function regulate dynamics at these sites. To accomplish functions, zyxin recruits VASP cellular via proline-rich binding near zyxin’s amino terminus. While the prevailing thought has been that simply acts as a scaffold protein for binding, identification LIM domain-VASP interaction could complicate...
Many of the diseases that plague society today are driven by a loss protein quality. One method to quantify quality is measure folding stability (PFS). Here, we present novel mass spectrometry (MS)-based approach for PFS measurement, iodination assay (IPSA). IPSA quantifies tracking surface-accessibility differences tyrosine, histidine, methionine, and cysteine under denaturing conditions. Relative current methods, increases coverage granularity track changes along its sequence. To our...
Abstract TELSAM crystallization promises to become a revolutionary tool for the facile of proteins. can increase rate and form crystals at low protein concentrations without direct contact between polymers and, in some cases, with very minimal crystal contacts overall (Nawarathnage et al ., 2022). To further understand characterize TELSAM-mediated crystallization, we sought requirements composition linker fused target protein. We evaluated four different linkers Ala-Ala, Ala-Val, Thr-Val,...
TELSAM-fusion crystallization has the potential to become a revolutionary tool for facile of proteins. TELSAM fusion can increase rate and enable at low protein concentrations, in some cases with minimal crystal contacts [Nawarathnage et al. (2022), Open Biol. 12 , 210271]. Here, requirements linker composition between 1TEL fused CMG2 vWa domain were investigated. Ala-Ala, Ala-Val, Thr-Val Thr-Thr linkers evaluated, comparing metrics propensity order. The effect on removing or retaining...
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