Chemistry and Biochemistry Faculty Works

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Open Access
Ancient hybridization leads to the repeated evolution of red flowers across a monkeyflower radiation
(Oxford Academic, 2023-06-05) Short, Aidan W.; Streisfeld, Matthew A.
The reuse of old genetic variation can promote rapid diversification in evolutionary radiations, but in most cases, the historical events underlying this divergence are not known. For example, ancient hybridization can generate new combinations of alleles that sort into descendant lineages, potentially providing the raw material to initiate divergence. In the Mimulus aurantiacus species complex, there is evidence for widespread gene flow among members of this radiation. In addition, allelic variation in the MaMyb2 gene is responsible for differences in flower color between the closely related ecotypes of subspecies puniceus, contributing to reproductive isolation by pollinators. Previous work suggested that MaMyb2 was introgressed into the red-flowered ecotype of puniceus. However, additional taxa within the radiation have independently evolved red flowers from their yellow-flowered ancestors, raising the possibility that this introgression had a more ancient origin. In this study, we used repeated tests of admixture from whole-genome sequence data across this diverse radiation to demonstrate that there has been both ancient and recurrent hybridization in this group. However, most of the signal of this ancient introgression has been removed due to selection, suggesting that widespread barriers to gene flow are in place between taxa. Yet, a roughly 30 kb region that contains the MaMyb2 gene is currently shared only among the red-flowered taxa. Patterns of admixture, sequence divergence, and extended haplotype homozygosity across this region confirm a history of ancient hybridization, where functional variants have been preserved due to positive selection in red-flowered taxa but lost in their yellow-flowered counterparts. The results of this study reveal that selection against gene flow can reduce genomic signatures of ancient hybridization, but that historical introgression can provide essential genetic variation that facilitates the repeated evolution of phenotypic traits between lineages.
Open Access
Molecular dissection of PI3Kβ synergistic activation by receptor tyrosine kinases, GβGγ, and Rho-family GTPases
(bioRxiv, 2023-05-01) Duewell, Benjamin R.; Wilson, Naomi E.; Bailey, Gabriela M.; Peabody, Sarah E.; Hansen, Scott D.
The class 1A phosphoinositide 3-kinase (PI3K) beta (PI3Kβ) is functionally unique in the ability to integrate signals derived from receptor tyrosine kinases (RTKs), heterotrimeric guanine nucleotidebinding protein (G-protein)-coupled receptors (GPCRs), and Rho-family GTPases. The mechanism by which PI3Kβ prioritizes interactions with various membrane tethered signaling inputs, however, remains unclear. Previous experiments have not been able to elucidate whether interactions with membranetethered proteins primarily control PI3Kβ localization versus directly modulate lipid kinase activity. To address this gap in our understanding of PI3Kβ regulation, we established an assay to directly visualize and decipher how three binding interactions regulate PI3Kβ when presented to the kinase in a biologically relevant configuration on supported lipid bilayers. Using single molecule Total Internal Reflection Fluorescence (TIRF) Microscopy, we determined the mechanism controlling membrane localization of PI3Kβ, prioritization of signaling inputs, and lipid kinase activation. We find that auto-inhibited PI3Kβ must first cooperatively engage a single RTK-derived tyrosine phosphorylated (pY) peptide before it can engage either GβGγ or Rac1(GTP). Although pY peptides strongly localize PI3Kβ to membranes, they only modestly stimulate lipid kinase activity. In the presence of either pY/GβGγ or pY/Rac1(GTP), PI3Kβ activity is dramatically enhanced beyond what can be explained by the increase in membrane avidity for these complexes. Instead, PI3Kβ is synergistically activated by pY/GβGγ and pY/Rac1(GTP) through a mechanism of allosteric regulation.
Open Access
Ligand field tuning of d-orbital energies in MOF clusters
(Research Square, 2023-04-12) Diamond, Brian; Payne, Lillian; Hendon, Christopher
Linker functionalization is a common route used to affect the electronic and catalytic properties of metal-organic frameworks. By either pre- or post-synthetically installing linkages with differing linker moieties the band gap, workfunction, and exciton lifetimes have been shown to be affected. One overlooked aspect of linker functionalization, however, has been the impact on the metal dorbital energies to which they are bound. The ligand field differences should result in substantial changes in d-splitting. In this study we use DFT to study the energetics of d-orbital energy tuning as a function of linker chemistry. We offer a general descriptor, linker pKa, as a tool to predict the resultant d-splitting in MOFs. Our calculations reveal that simple functionalizations can affect the d-energies by up to 2 eV and illustrate the significance of this band modularity using four archetypal MOFs: UiO-66, MIL-125, ZIF-8, and MOF-5. Together, we show that linker functionalization dramatically affects d-energies in MOF clusters and highlight that linker functionalization is a useful route for fine-tuning band edges centered on the metals, rather than linkers themselves.
Open Access
Mendeleev eponyms in the epoch of educational ethnocentrism
(Scientia Socialis, 2023) Slabin, Uladzimir
Eponymous terms play an important role in STEM education. This research focuses on the current state of Mendeleev eponyms in the context of education and ethnocentrism, addressing their usage in various languages, their educational value, cases of questioned priority and copyright violation in Mendeleev major eponyms–periodic table and periodic system. 106 chemistry textbooks in 4 languages including Soviet-time and current Russian textbooks were perused to identify and trace Mendeleev eponyms over 1924-2016. Advanced Google Search with queries in Belarusian, English, Latvian, Polish, Russian, and Ukrainian was conducted to evaluate online presence of eponyms “Mendeleev periodic table” and “Mendeleev periodic system.” It was found that while Mendeleev eponyms occur generously on the Internet, periodic table and system with Mendeleev’s name attached are seldom used on non-Russian webpages. Most Mendeleev eponyms were made up in the USSR and remain popular and Russia, which can be explained within the framework of ethnocentrism as a ruling tendency. Recognizing Mendeleev’s priority, Flinn and Ross’s periodic systems can be considered plagiarized; a few factors might favor their emergence, but ethnocentrism is unlikely to be one of them. Mendeleev eponyms remain valuable assets for science education, acting as shortcuts to the history of science and actualizing interdisciplinary connections.
Open Access
Aging and sperm signals alter DNA break formation and repair in the C. elegans germline
(PLOS, 2022-11-07) Toraason, Erik; Adler, Victoria L.; Libuda, Diana E.
Female reproductive aging is associated with decreased oocyte quality and fertility. The nematode Caenorhabditis elegans is a powerful system for understanding the biology of aging and exhibits age-related reproductive defects that are analogous to those observed in many mammals, including dysregulation of DNA repair. C. elegans germline function is influenced simultaneously by both reproductive aging and signals triggered by limited supplies of sperm, which are depleted over chronological time. To delineate the causes of DNA repair defects in aged C. elegans germlines, we assessed both DNA double strand break (DSB) induction and repair during meiotic prophase I progression in aged germlines which were depleted of self-sperm, mated, or never exposed to sperm. We find that germline DSB induction is dramatically reduced only in hermaphrodites which have exhausted their endogenous sperm, suggesting that a signal due specifically to sperm depletion downregulates DSB formation. We also find that DSB repair is delayed in aged germlines regardless of whether hermaphrodites had either a reduction in sperm supply or an inability to endogenously produce sperm. These results demonstrate that in contrast to DSB induction, DSB repair defects are a feature of C. elegans reproductive aging independent of sperm presence. Finally, we demonstrate that the E2 ubiquitin-conjugating enzyme variant UEV-2 is required for efficient DSB repair specifically in young germlines, implicating UEV-2 in the regulation of DNA repair during reproductive aging. In summary, our study demonstrates that DNA repair defects are a feature of C. elegans reproductive aging and uncovers parallel mechanisms regulating efficient DSB formation in the germline.
Open Access
Comparison of fractal and grid electrodes for studying the effects of spatial confinement on dissociated retinal neuronal and glial behavior
(Nature, 2022-10-20) Moslehi, Saba; Rowland, Conor; Smith, Julian H.; Griffiths, Willem; Watterson, William J.; Niell, Cristopher M.; Alemán, Benjamín J.; Perez, Maria-Thereza; Taylor, Richard P.
Understanding the impact of the geometry and material composition of electrodes on the survival and behavior of retinal cells is of importance for both fundamental cell studies and neuromodulation applications. We investigate how dissociated retinal cells from C57BL/6J mice interact with electrodes made of vertically-aligned carbon nanotubes grown on silicon dioxide substrates. We compare electrodes with different degrees of spatial confinement, specifically fractal and grid electrodes featuring connected and disconnected gaps between the electrodes, respectively. For both electrodes, we find that neuron processes predominantly accumulate on the electrode rather than the gap surfaces and that this behavior is strongest for the grid electrodes. However, the ‘closed’ character of the grid electrode gaps inhibits glia from covering the gap surfaces. This lack of glial coverage for the grids is expected to have long-term detrimental effects on neuronal survival and electrical activity. In contrast, the interconnected gaps within the fractal electrodes promote glial coverage. We describe the differing cell responses to the two electrodes and hypothesize that there is an optimal geometry that maximizes the positive response of both neurons and glia when interacting with electrodes.
Open Access
Anomalous Dynamics in Macromolecular Liquids
(MDPI, 2022-02-22) Guenza, Marina G.
Macromolecular liquids display short-time anomalous behaviors in disagreement with conventional single-molecule mean-field theories. In this study, we analyze the behavior of the simplest but most realistic macromolecular system that displays anomalous dynamics, i.e., a melt of short homopolymer chains, starting from molecular dynamics simulation trajectories. Our study sheds some light on the microscopic molecular mechanisms responsible for the observed anomalous behavior. The relevance of the correlation hole, a unique property of polymer liquids, in relation to the observed subdiffusive dynamics, naturally emerges from the analysis of the van Hove distribution functions and other properties.
Open Access
Dinucleotides as simple models of the base stacking-unstacking component of DNA 'breathing' mechanisms
(Oxford Academic, 2021-01) Beyerle, Eric R.; Dinpajooh, Mohammadhasan; Ji, Huiying; von Hippel, Peter H.; Marcus, Andrew H.; Guenza, Marina G.
Regulatory protein access to the DNA duplex 'interior' depends on local DNA 'breathing' fluctuations, and the most fundamental of these are thermally-driven base stacking-unstacking interactions. The smallest DNA unit that can undergo such transitions is the dinucleotide, whose structural and dynamic properties are dominated by stacking, while the ion condensation, cooperative stacking and inter-base hydrogen-bonding present in duplex DNA are not involved. We use dApdA to study stacking-unstacking at the dinucleotide level because the fluctuations observed are likely to resemble those of larger DNA molecules, but in the absence of constraints introduced by cooperativity are likely to be more pronounced, and thus more accessible to measurement. We study these fluctuations with a combination of Molecular Dynamics simulations on the microsecond timescale and Markov State Model analyses, and validate our results by calculations of circular dichroism (CD) spectra, with results that agree well with the experimental spectra. Our analyses show that the CD spectrum of dApdA is defined by two distinct chiral conformations that correspond, respectively, to a Watson-Crick form and a hybrid form with one base in a Hoogsteen configuration. We find also that ionic structure and water orientation around dApdA play important roles in controlling its breathing fluctuations.
Open Access
Mapping DNA conformations and interactions within the binding cleft of bacteriophage T4 single-stranded DNA binding protein (gp32) at single nucleotide resolution
(Oxford University Press, 2020-12-24) Camel, Benjamin R.; Jose, Davis; Meze, Katarina; Dang, Anson; von Hippel, Peter H.
In this study, we use single-stranded DNA (oligo-dT) lattices that have been position-specifically labeled with monomer or dimer 2-aminopurine (2-AP) probes to map the local interactions of the DNA bases with the nucleic acid binding cleft of gp32, the single-stranded binding (ssb) protein of bacteriophage T4. Three complementary spectroscopic approaches are used to characterize these local interactions of the probes with nearby nucleotide bases and amino acid residues at varying levels of effective protein binding cooperativity, as manipulated by changing lattice length. These include: (i) examining local quenching and enhancing effects on the fluorescence spectra of monomer 2-AP probes at each position within the cleft; (ii) using acrylamide as a dynamic-quenching additive to measure solvent access to monomer 2-AP probes at each ssDNA position; and (iii) employing circular dichroism spectra to characterize changes in exciton coupling within 2-AP dimer probes at specific ssDNA positions within the protein cleft. The results are interpreted in part by what we know about the topology of the binding cleft from crystallographic studies of the DNA binding domain of gp32 and provide additional insights into how gp32 can manipulate the ssDNA chain at various steps of DNA replication and other processes of genome expression.
Open Access
Evolving Stark Effect During Growth of Perovskite Nanocrystals Measured Using Transient Absorption
(Frontiers, 2020-10) Sadighian, James C.; Wilson, Kelly S.; Crawford, Michael L.; Wong, Cathy Y.
Methylammonium lead triiodide (MAPbI3) nanocrystals (NCs) are emerging materials for a range of optoelectronic applications. Photophysical characterization is typically limited to structurally stable NCs owing to the long timescales required for many spectroscopies, preventing the accurate measurement of NCs during growth. This is a particular challenge for non-linear spectroscopies such as transient absorption. Here we report on the use of a novel single-shot transient absorption (SSTA) spectrometer to study MAPbI3 NCs as they grow. Comparing the transient spectra to derivatives of the linear absorbance reveals that photogenerated charge carriers become localized at surface trap states during NC growth, inducing a TA lineshape characteristic of the Stark effect. Observation of this Stark signal shows that the contribution of trapped carriers to the TA signal declines as growth continues, supporting a growth mechanism with increased surface ligation toward the end of NC growth. This work opens the door to the application of time-resolved spectroscopies to NCs in situ, during their synthesis, to provide greater insight into their growth mechanisms and the evolution of their photophysical properties.
Open Access
Quinoidal diindenothienoacenes: synthesis and properties of new functional organic materials
(Royal Society of Chemistry, 2014-06-16) Rudebusch, Gabriel; Fix, Aaron; Henthorn, Hilary; Vonnegut, Chris; Zakharov, Lev; Haley, Michael
We report the preparation and characterization of a new class of quinoidal thienoacenes. The synthetic route is efficient, high-yielding and scalable with the potential for further functionalization. Single crystal X-ray diffraction reveals that, as size increases, the molecules pack in progressively closer 1D arrangements. The title compounds are shown to have amphoteric redox behaviour by cyclic voltammetry. The anion radicals are studied by EPR spectrometry and by computations. The electron-accepting nature, NIR absorption and the low-lying LUMO energies (ca. −4.0 eV) allude to potential use in materials applications.
Open Access
Synthesis and properties of fully-conjugated indacenedithiophenes
(Chemical Science, 2013-12-10) Young, Brian; Chase, Daniel Tyler, 1983-; Marshall, Jonathan; Vonnegut, Chris; Zakharov, Lev; Haley, Michael
The synthesis and characterization of four fully-conjugated indacenedithiophenes (IDTs) are disclosed. In contrast to anthradithiophenes, regioselective synthesis of both syn and anti isomers is readily achieved. Thiophene fusion imparts increased paratropic character on the central indacene core as predicted by DFT calculations and confirmed by 1H NMR spectroscopy. IDTs exhibit red-shifted absorbance maxima with respect to their all-carbon analogues and undergo two-electron reduction and one-electron oxidation.
Open Access
Beyond Information: Exploring Patients’ Preferences
(American Medical Association, 2009) Epstein, Ronald; Peters, Ellen
The Institute of Medicine considers patient-centered care (“care that is respectful of and responsive to individual patient preferences, needs and values” 1(p6)) to be a foundation of high-quality health care, along with effectiveness, safety, efficiency, timeliness, and equity. Patient-centered care is empirically based and promotes respect and patient autonomy; it is considered an end in itself, not merely a means to achieve other health outcomes.2 Two parallel efforts have furthered patient-centered care. Shared decision making promotes defining problems, presenting options, and providing high-quality information so patients can participate more actively in care.3 Patient-centered communication promotes healing relationships that elicit and consider patients’ perspectives and understand patients as persons. 2 Both approaches assume that patients can articulate preferences based on stable guiding principles or values. While this may be true in straightforward situations, in novel, unanticipated, and emotionally charged situations, preferences may not be elicited as much as they are constructed—shaped by how information is presented and by the opinions of family, friends, and the media. This Commentary explores how physicians might reconcile the imperative to provide patient-centered care with the complex ways in which clinicians and patients construct preferences.
Open Access
Tetracarbonylbis(η5-cyclopentadienyl)bis[(dec-9-en-1-yl)diphenylphosphine]dimolybdenum(0)(Mo--Mo) tetrahydrofuran disolvate
(International Union of Crystallography, 2009) Schultz, Ginger; Zakharov, Lev; Tyler, David R.
The asymmetric unit of the title compound, [Mo2( 5- C5H5)2(C22H29P)2(CO)4] 2C4H8O, contains two half-molecules of the organometallic species and two solvent molecules. Both organometallic molecules are completed by crystallographic inversion symmetry, yielding dimeric units with Mo—Mo single-bond lengths of 3.2703 (6) and 3.2548 (6) A ˚ . Each Mo atom is also coordinated by an 5- cyclopentdienyl ligand, two carbonyl ligands, and a (dec-9-en- 1-yl)diphenylphosphine ligand.
Open Access
Prion protein expression in Chinese hamster ovary cells using a glutamine synthetase selection and amplification system
(Protein Engineering, 1997-09-05) Blochberger, Thomas C.; Cooper, Carl; Peretz, David; Tatzelt, Jorg
Syrian hamster prion protein (PrPc) and a truncated Syrian hamster prion protein lacking the glycosylphosphatidylinositol (GPI) anchor C-terminal signal sequence (GPI~) were expressed in Chinese hamster ovary cells using a glutamine synthetase selection and amplification system. The CHO cell clones expressing the GPI" PrP secreted the majority of the protein into the media, whereas most of the PrP produced by clones expressing the full-length protein with the GPI anchor was located on the cell surface, as demonstrated by its release upon treatment with phosphatidylinositol-specific phospholipase C (PIPLC). A cell clone that expressed the highest levels of full length PrP was subcloned to obtain clone 30C3-1. PrP from clone 30C3-1 was shown to be sensitive to proteolysis by proteinase K and to react with monoclonal and polyclonal antibodies that recognize native PrPc. The recombinant PrP migrated as a diffuse band of 19-40 kDa but removal of the N-linked oligosaccharides with peptide N-glycosidase F (PNGase F) revealed three protein species of 19, 17 and 15 kDa. The 19 kDa band corresponding to deglycosylated full-length PrP was quantified and found to be expressed at a level - 14-fold higher than that of PrPc found in Syrian hamster brain.
Open Access
Prion protein expression in Chinese hamster ovary cells using a glutamine synthetase selection and amplification system
(Protein Engineering, 1997) Blochberger, Thomas C.; Cooper, Carl; Peretz, David; Tatzelt, Jorg; Griffith, O. H.; Baldwin, Michael A.; Prusiner, Stanley B.
Open Access
Prion protein (PrP) synthetic peptides induce cellular PrP to acquire properties of the scrapie isoform
(Proceedings of the National Academy of Sciences, 1995-11) Kaneko, K.; Peretz, David; Pan, K. K.; Blochberger, Thomas C.; Wille, H.; Gabizon, R.; Griffith, O. H.; Cohen, F. E.; Baldwin, Michael A.; Prusiner, Stanley B.
Conversion of the cellular isoform of prion protein (PrPc) into the scrapie isoform (PrPSc) involves an increase in the /3-sheet content, diminished solubility, and resistance to proteolytic digestion. Transgenetic studies argue that PrPc and PrPSc form a complex during PrPScformation; thus, synthetic PrP peptides, which mimic the conformational pluralism of PrP, were mixed with PrPc to determine whether its properties were altered. Peptides encompassing two a-helical domains of PrP when mixed with PrPc produced a complex that displayed many properties of PrPSc. The PrPcpeptide complex formed fibrous aggregates and up to 65% of complexed PrPc sedimented at 100,000 x g for 1 h, whereas PrPc alone did not. These complexes were resistant to pro teolytic digestion and displayed a high /3-sheet content. Un expectedly, the peptide in a /3-sheet conformation did not form the complex, whereas the random coil did. Addition of 2% Sarkosyl disrupted the complex and rendered PrPc sensitive to protease digestion. While the pathogenic AllTV mutation increased the efficacy of complex formation, anti-PrP mono clonal antibody prevented interaction between PrPc and pep tides. Our findings in concert with transgenetic investigations argue that PrPc interacts with PrPSc through a domain that contains the first two putative a-helices. Whether PrPc-peptide complexes possess prion infectivity as determined by bioassays remains to be established.
Open Access
Electron optical benches for in-line and branched systems. A new bench designed for mirror-based aberration correction and low energy electron microscopy
(Review of Scientific Instruments, 1994-10) Skoczylas, W. P.; Rempfer, G. F.; Griffith, O. H.
A review of electron optical bench literature is presented, and the designs of two optical benches used by the authors are described. One bench was designed for testing individual electrostatic electron lenses and in-line optical systems, for example, emission electron microscopes and transmission electron microscopes. It has been in operation for many years. The second electron optical bench is new. It is a branched system designed for several purposes: to study correction of spherical and chromatic aberration with an electron mirror, and to gain experience with low energy electron microscopy (LEEM) optics. The alignment of the electron optical support structure is independent of the vacuum housing, and the bench is designed to be operated either horizontally or vertically. As a demonstration of the performance of the new bench in the horizontal mode, a test pattern on a silicon surface was imaged with LEEM optics.
Open Access
Phorbol ester-induced actin cytoskeletal reorganization requires a heavy metal ion, probably Zn2+.
(Cell Regulation, 1991-12) Hedberg, K. K.; Birrell, G. B.; Griffith, O. H.
The cell-permeant heavy metal chelator N,N,N',Ntetrakis( 2-pyridylmethyl)ethylenediamine (TPEN) was found to counteract phorbol ester-induced actin reorganization in PTK2 and Swiss 3T3 cells. By us ing fluorescence and the higher resolution tech nique of photoelectron microscopy to monitor actin patterns, 15-min pretreatment with 25-50 pM TPEN was found to dramatically reduce actin alterations resulting from subsequent phorbol ester treatment in PTK2 cells. Similar results were obtained with Swiss 3T3 cells using 50 mM TPEN for 1.5 h. Phorbol ester-induced actin alterations are thought to de pend on activation of protein kinase C (PKC). In contrast to the phorbol ester effect, the PKC-independent actin cytoskeletal disruption caused by staurosporine and cytochalasin B was unaffected by TPEN pretreatment. TPEN did not block phorbol ester-induced activation of PKC in Swiss 3T3 cells, as observed by the phosphorylation of the 80K PKC substrate protein (MARCKS protein). TPEN also did not inhibit partially purified PKC from Swiss 3T3 cells in an in vitro PKC-specific commercial assay. To establish that the effect of TPEN is the removal of metal ions and not some other nonspecific effect of TPEN, a series of transition metal ions was added at the end of the TPEN pretreatment. The results indicate that the transient but dramatic phorbol ester-induced reorganization of the actin cytoskeleton in cultured cells depends on an inter action of PKC with a heavy metal, probably zinc.
Open Access
Phorbol ester-induced actin cytoskeletal reorganization requires a heavy metal ion
(Cell Regulation, 1991-12) Hedberg, Karen K.; Birrell, G. B.; Griffith, O. H.