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Journal Article
Dooner, HK, He L.  2014.  Polarized gene conversion at the bz locus of maize.. Proc Natl Acad Sci USA. 111(38):13918-23. Abstract
Nucleotide diversity is greater in maize than in most organisms studied to date, so allelic pairs in a hybrid tend to be highly polymorphic. Most recombination events between such pairs of maize polymorphic alleles are crossovers. However, intragenic recombination events not associated with flanking marker exchange, corresponding to noncrossover gene conversions, predominate between alleles derived from the same progenitor. In these dimorphic heterozygotes, the two alleles differ only at the two mutant sites between which recombination is being measured. To investigate whether gene conversion at the bz locus is polarized, two large diallel crossing matrices involving mutant sites spread across the bz gene were performed and more than 2,500 intragenic recombinants were scored. In both diallels, around 90% of recombinants could be accounted for by gene conversion. Furthermore, conversion exhibited a striking polarity, with sites located within 150 bp of the start and stop codons converting more frequently than sites located in the middle of the gene. The implications of these findings are discussed with reference to recent data from genome-wide studies in other plants.
Kumaraswamy, GK, Guerra T, Qian X, Zhang S, Bryant DA, Dismukes GC.  2013.  Reprogramming the glycolytic pathway for increased hydrogen production in cyanobacteria: metabolic engineering of NAD+-dependent GAPDH. Energy Environ. Sci.. 6:3722-3731. AbstractWebsite
Catabolism of glycogen stored by cyanobacteria occurs during anaerobic auto-fermentation and produces a range of C1–C3 fermentation products and hydrogen via hydrogenase. We investigated both augmenting and rerouting this carbon catabolism by engineering the glycolysis pathway at the NAD+-dependent glyceraldehyde-3-phosphate dehydrogenase (GAPDH-1), its major regulation site at the nexus of two pathways (Oxidative Pentose Phosphate pathway, OPP, and glycolysis/gluconeogenesis). Null (gap1::aphII) and overexpression (gap1+) strains of Synechococcus sp. strain PCC 7002 were constructed in order to produce more NADPH (via rerouting carbon through OPP) and more NADH (via opening the glycolytic bottleneck), respectively. For gap1::aphII quantitative analyses after four days of dark auto-fermentation showed undiminished glycogen catabolism rate, significant increases of intracellular metabolites in both OPP and upper-glycolysis, decrease in lower-glycolysis intermediates, 5.7-fold increase in NADPH, 2.3-fold increase in hydrogen and 1.25-fold increase in CO2vs. wild type (WT). These changes demonstrate the expected outcome of redirection of carbon catabolism through the OPP pathway with significant stimulation of OPP product yields. The gap1+ strain exhibits a large 17% increase in accumulation of glycogen during the prior photoautotrophic growth stage (gluconeogenesis), in parallel with a 2-fold increase in the total [NAD+ + NADH] pool, foreshadowing an increased catabolic capacity. Indeed, the rate of glycogen catabolism during subsequent dark auto-fermentation increased significantly (58%) vs. WT, resulting in increases in both NADH (4.0-fold) and NADPH (2.9-fold) pools, and terminal fermentation products, hydrogen (3.0-fold) D-lactate (2.3-fold) and acetate (1.4-fold). The overall energy conversion yield over four days from catabolized glycogen to hydrogen increased from 0.6 mole of hydrogen per mole of glucose (WT) to 1.4 (gap1::aphII) and 1.1 (gap1+) under headspace accumulation conditions (without hydrogen milking). These findings demonstrate the significant potential of metabolic engineering for redirecting carbon pathways for carbohydrate catabolism and hydrogen production in cyanobacteria.
Gates, C, Ananyev GM, Dismukes C.  2016.  The strontium inorganic mutant of the water oxidizing center (CaMn4O5) of PSII improves WOC efficiency but slows electron flux through the terminal acceptors.. Biochim Biophys Acta.. 1857(9):1550-1560. Abstract
Herein we extend prior studies of biosynthetic strontium replacement of calcium in PSII-WOC core particles to characterize whole cells. Previous studies of Thermosynechococcus elongatus found a lower rate of light-saturated O2 from isolated PSII-WOC(Sr) cores and 5–8 × slower rate of oxygen release. We find similar properties in whole cells, and show it is due to a 20% larger Arrhenius activation barrier for O2 evolution. Cellular adaptation to the sluggish PSII-WOC(Sr) cycle occurs in which flux through the QAQB acceptor gate becomes limiting for turnover rate in vivo. Benzoquinone derivatives that bind to QB site remove this kinetic chokepoint yielding 31% greater O2 quantum yield (QY) of PSII-WOC(Sr) vs. PSII-WOC(Ca). QY and efficiency of the WOC(Sr) catalytic cycle are greatly improved at low light flux, due to fewer misses and backward transitions and 3-fold longer lifetime of the unstable S3 state, attributed to greater thermodynamic stabilization of the WOC(Sr) relative to the photoactive tyrosine YZ. More linear and less cyclic electron flow through PSII occurs per PSII-WOC(Sr). The organismal response to the more active PSII centers in Sr-grown cells at 45 °C is to lower the number of active PSII-WOC per Chl, producing comparable oxygen and energy per cell. We conclude that redox and protonic energy fluxes created by PSII are primary determinants for optimal growth rate of T. elongatus. We further conclude that the (Sr-favored) intermediate-spin S = 5/2 form of the S2 state is the active form in the catalytic cycle relative to the low-spin S = 1/2 form.
Deibert, BJ, Zhang J, Smith PF, Chapman KW, Rangan S, Banerjee D, Tan K, Wang H, Pasguale N, Chen F et al..  2015.  Surface and Structural Investigation of a MnOx Birnessite-Type Water Oxidation Catalyst Formed under Photocatalytic Conditions. Chemistry. 21(40):14218-14228. Abstract
Catalytically active MnOx species have been reported to form in situ from various Mn-complexes during electrocatalytic and solution-based water oxidation when employing cerium(IV) ammonium ammonium nitrate (CAN) oxidant as a sacrificial reagent. The full structural characterization of these oxides may be complicated by the presence of support material and lack of a pure bulk phase. For the first time, we show that highly active MnOx catalysts form without supports in situ under photocatalytic conditions. Our most active (4)MnOx catalyst (∼0.84 mmol O2  mol Mn(-1) s(-1)) forms from a Mn4O4 bearing a metal-organic framework. (4)MnOx is characterized by pair distribution function analysis (PDF), Raman spectroscopy, and HR-TEM as a disordered, layered Mn-oxide with high surface area (216 m(2) g(-1)) and small regions of crystallinity and layer flexibility. In contrast, the (S)MnOx formed from Mn(2+) salt gives an amorphous species of lower surface area (80 m(2) g(-1)) and lower activity (∼0.15 mmol O2  mol Mn(-1) s(-1)). We compare these catalysts to crystalline hexagonal birnessite, which activates under the same conditions. Full deconvolution of the XPS Mn2p3/2 core levels detects enriched Mn(3+) and Mn(2+) content on the surfaces, which indicates possible disproportionation/comproportionation surface equilibria.
Li, Y, Harris L, Dooner HK.  2013.  TED, an autonomous and rare maize transposon of the mutator superfamily with a high gametophytic excision frequency.. The Plant cell. 25(9):3251-65. Abstract
Mutator (Mu) elements, one of the most diverse superfamilies of DNA transposons, are found in all eukaryotic kingdoms, but are particularly numerous in plants. Most of the present knowledge on the transposition behavior of this superfamily comes from studies of the maize (Zea mays) Mu elements, whose transposition is mediated by the autonomous Mutator-Don Robertson (MuDR) element. Here, we describe the maize element TED (for Transposon Ellen Dempsey), an autonomous cousin that differs significantly from MuDR. Element excision and reinsertion appear to require both proteins encoded by MuDR, but only the single protein encoded by TED. Germinal excisions, rare with MuDR, are common with TED, but arise in one of the mitotic divisions of the gametophyte, rather than at meiosis. Instead, transposition-deficient elements arise at meiosis, suggesting that the double-strand breaks produced by element excision are repaired differently in mitosis and meiosis. Unlike MuDR, TED is a very low-copy transposon whose number and activity do not undergo dramatic changes upon inbreeding or outcrossing. Like MuDR, TED transposes mostly to unlinked sites and can form circular transposition products. Sequences closer to TED than to MuDR were detected only in the grasses, suggesting a rather recent evolutionary split from a common ancestor.
Hawkins, JS, Delgado V, Feng L, Carlise M, Dooner HK, Bennetzen JL.  2014.  Variation in allelic expression associated with a recombination hotspot in Zea mays.. The Plant Journal, DOI: 10.1111/tpj.12537. Abstract
Gene expression is a complex process, requiring precise spatial and temporal regulation of transcription factor activity; however, modifications of individual cis- and trans-acting modules can be molded by natural selection to create a sizeable number of novel phenotypes. Results from decades of research indicate that developmental and phenotypic divergence among eukaryotic organisms is driven primarily by variation in levels of gene expression that are dictated by mutations either in structural or regulatory regions of genes. The relative contributions and interplay of cis- and trans-acting regulatory factors to this evolutionary process, however, remain poorly understood. Analysis of 8 genes in the Bz1-Sh1 interval of maize indicates significant allele-specific expression biases in at least one tissue for all genes, ranging from 1.3-fold to 36-fold. All detected effects were cis-regulatory in nature, although genetic background may also influence the level of expression bias and tissue specificity for some allelic combinations. Most allelic pairs exhibited the same direction and approximate intensity of bias across all four tissues; however, a subset of allelic pairs show alternating dominance across different tissue types or variation in the degree of bias in different tissues. In addition, the genes showing the most striking levels of allelic bias co-localize with a previously described recombination hotspot in this region, suggesting a naturally occurring genetic mechanism for creating regulatory variability for a subset of plant genes that may ultimately lead to evolutionary diversification.
Smith, PF, Hunt L, Laursen AB, Sagar V, Kaushik S, Calvinho KU, Marotta G, Mosconi E, De Angelis F, Dismukes GC.  2015.  Water Oxidation by the [Co4O4(OAc)4(py)4](+) Cubium is Initiated by OH(-) Addition.. J Am Chem Soc. 137(49):15460-15468. Abstract
The cobalt cubium Co4O4(OAc)4(py)4(ClO4) (1A(+)) containing the mixed valence [Co4O4](5+) core is shown by multiple spectroscopic methods to react with hydroxide (OH(-)) but not with water molecules to produce O2. The yield of reaction products is stoichiometric (>99.5%): 41A(+) + 4OH(-) → O2 + 2H2O + 41A. By contrast, the structurally homologous cubium Co4O4(trans-OAc)2(bpy)4(ClO4)3, 1B(ClO4)3, produces no O2. EPR/NMR spectroscopies show clean conversion to cubane 1A during O2 evolution with no Co(2+) or Co3O4 side products. Mass spectrometry of the reaction between isotopically labeled μ-(16)O(bridging-oxo) 1A(+) and (18)O-bicarbonate/water shows (1) no exchange of (18)O into the bridging oxos of 1A(+), and (2) (36)O2 is the major product, thus requiring two OH(-) in the reactive intermediate. DFT calculations of solvated intermediates suggest that addition of two OH(-) to 1A(+) via OH(-) insertion into Co-OAc bonds is energetically favored, followed by outer-sphere oxidation to intermediate [1A(OH)2](0). The absence of O2 production by cubium 1B(3+) indicates the reactive intermediate derived from 1A(+) requires gem-1,1-dihydoxo stereochemistry to perform O-O bond formation. Outer-sphere oxidation of this intermediate by 2 equiv of 1A(+) accounts for the final stoichiometry. Collectively, these results and recent literature (Faraday Discuss., doi:10.1039/C5FD00076A and J. Am. Chem. Soc. 2015, 137, 12865-12872) validate the [Co4O4](4+/5+) cubane core as an intrinsic catalyst for oxidation of hydroxide by an inner-sphere mechanism.