Biochemistry, Biophysics, and Structural Biology Commons^™

Mutational Analysis Of The Nitrogenase Carbon Monoxide Protective Protein Cown Reveals That A Conserved C‑Terminal Glutamic Acid Residue Is Necessary For Its Activity, Dustin L. Willard, Joshuah J. Arellano, Mitch Underdahl, Terrence M. Lee, Avinash S. Ramaswamy, Gabriella Fumes, Agatha Kliman, Emily Y. Wong, Cedric P. Owens

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Nitrogenase is the only enzyme that catalyzes the reduction of nitrogen gas into ammonia. Nitrogenase is tightly inhibited by the environmental gas carbon monoxide (CO). Many nitrogen fixing bacteria protect nitrogenase from CO inhibition using the protective protein CowN. This work demonstrates that a conserved glutamic acid residue near the C-terminus of Gluconacetobacter diazotrophicus CowN is necessary for its function. Mutation of the glutamic acid residue abolishes both CowN’s protection against CO inhibition and the ability of CowN to bind to nitrogenase. In contrast, a conserved C-terminal cysteine residue is not important for CO protection by CowN. Overall, this work …

Arginine Methylation Of The Pgc-1Α C‑Terminus Is Temperature- Dependent, Meryl Mendoz, Mariel Mendoza, Tiffany Lubrino, Sidney Briski, Immaculeta Osuji, Janielle Cuala, Brendan Ly, Ivan Ocegueda, Harvey Peralta, Benjamin A. Garcia, Cecilia Zurita-Lopez

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

We set out to determine whether the C-terminus (amino acids 481–798) of peroxisome proliferator-activated receptor gamma coactivator-1 alpha (PGC-1α, UniProt Q9UBK2), a regulatory metabolic protein involved in mitochondrial biogenesis, and respiration, is an arginine methyltransferase substrate. Arginine methylation by protein arginine methyltransferases (PRMTs) alters protein function and thus contributes to various cellular processes. In addition to confirming methylation of the C-terminus by PRMT1 as described in the literature, we have identified methylation by another member of the PRMT family, PRMT7. We performed in vitro methylation reactions using recombinant mammalian PRMT7 and PRMT1 at 37, 30, 21, 18, and 4 °C. …

Evaluating Alternative Ebullition Models For Predicting Peatland Methane Emission And Its Pathways Via Data–Model Fusion, Shuang Ma, Lifen Jiang, Rachel M. Wilson, Jeff P. Chanton, Scott Bridgham, Shuli Niu, Colleen M. Iversen, Avni Malhotra, Jiang Jiang, Xingjie Lu, Yuanyuan Huang, Jason Keller, Xiaofeng Xu, Daniel M. Ricciuto, Paul J. Hanson, Yiqi Luo

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Understanding the dynamics of peatland methane (CH4) emissions and quantifying sources of uncertainty in estimating peatland CH4 emissions are critical for mitigating climate change. The relative contributions of CH4 emission pathways through ebullition, plant-mediated transport, and diffusion, together with their different transport rates and vulnerability to oxidation, determine the quantity of CH4 to be oxidized before leaving the soil. Notwithstanding their importance, the relative contributions of the emission pathways are highly uncertain. In particular, the ebullition process is more uncertain and can lead to large uncertainties in modeled CH4 emissions. To improve model simulations of CH4 emission and its pathways, …

Evaluating Alternative Ebullition Models For Predicting Peatland Methane Emission And Its Pathways Via Data–Model Fusion, Shuang Ma, Lifen Jiang, Rachel M. Wilson, Jeff P. Chanton, Scott Bridgham, Shuli Niu, Colleen M. Iversen, Avni Malhotra, Jiang Jiang, Xingjie Lu, Jason Keller, Xiaofeng Xu, Daniel M. Ricciuto, Paul J. Hanson, Yiqi Luo

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Understanding the dynamics of peatland methane (CH₄) emissions and quantifying sources of uncertainty in estimating peatland CH₄ emissions are critical for mitigating climate change. The relative contributions of CH₄ emission pathways through ebullition, plant-mediated transport, and diffusion, together with their different transport rates and vulnerability to oxidation, determine the quantity of CH₄ to be oxidized before leaving the soil. Notwithstanding their importance, the relative contributions of the emission pathways are highly uncertain. In particular, the ebullition process is more uncertain and can lead to large uncertainties in modeled CH₄ emissions. To improve model simulations …

Escherichia Coli Alanyl-Trna Synthetase Maintains Proofreading Activity And Translational Accuracy Under Oxidative Stress, Arundhati Kavoor, Paul Kelly, Michael Ibba

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Aminoacyl-tRNA synthetases (aaRSs) are enzymes that synthesize aminoacyl-tRNAs to facilitate translation of the genetic code. Quality control by aaRS proofreading and other mechanisms maintains translational accuracy, which promotes cellular viability. Systematic disruption of proofreading, as recently demonstrated for alanyl-tRNA synthetase (AlaRS), leads to dysregulation of the proteome and reduced viability. Recent studies showed that environmental challenges such as exposure to reactive oxygen species can also alter aaRS synthetic and proofreading functions, prompting us to investigate if oxidation might positively or negatively affect AlaRS activity. We found that while oxidation leads to modification of several residues in Escherichia coli AlaRS, unlike …

Oxidation Alters The Architecture Of The Phenylalanyl-Trna Synthetase Editing Domain To Confer Hyperaccuracy, Pooja Srinivas, Rebecca E. Steiner, Ian J. Pavelich, Ricardo Guerrera-Ferreira, Puneet Juneja, Michael Ibba, Christine M. Dunham

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

High fidelity during protein synthesis is accomplished by aminoacyl-tRNA synthetases (aaRSs). These enzymes ligate an amino acid to a cognate tRNA and have proofreading and editing capabilities that ensure high fidelity. Phenylalanyl-tRNA synthetase (PheRS) preferentially ligates a phenylalanine to a tRNA^Phe over the chemically similar tyrosine, which differs from phenylalanine by a single hydroxyl group. In bacteria that undergo exposure to oxidative stress such as Salmonella enterica serovar Typhimurium, tyrosine isomer levels increase due to phenylalanine oxidation. Several residues are oxidized in PheRS and contribute to hyperactive editing, including against mischarged Tyr-tRNA^Phe, despite these oxidized residues not …

Cown Sustains Nitrogenase Turnover In The Presence Of The Inhibitor Carbon Monoxide, Michael S. Medina, Kevin O. Bretzing, Richard A. Aviles, Kiersten M. Chong, Alejandro Espinoza, Chloe Nicole G. Garcia, Benjamin B. Katz, Ruchita N. Kharwa, Andrea Hernandez, Justin L. Lee, Terrence M. Lee, Christine Lo Verde, Max W. Strul, Emily Y. Wong, Cedric P. Owens

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Nitrogenase is the only enzyme capable of catalyzing nitrogen fixation, the reduction of dinitrogen gas (N₂) to ammonia (NH₃). Nitrogenase is tightly inhibited by the environmental gas carbon monoxide (CO). Nitrogen-fixing bacteria rely on the protein CowN to grow in the presence of CO. However, the mechanism by which CowN operates is unknown. Here, we present the biochemical characterization of CowN and examine how CowN protects nitrogenase from CO. We determine that CowN interacts directly with nitrogenase and that CowN protection observes hyperbolic kinetics with respect to CowN concentration. At a CO concentration of 0.001 atm, …

Capsaicin Is A Negative Allosteric Modulator Of The 5-Ht3 Receptor, Eslam El Nebrisi, Tatiana Prytkova, Dietrich Ernst Lorke, Luke Howarth, Asma Hassan Alzaabi, Keun-Hang Susan Yang, Frank Christopher Howarth, Murat Oz

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

In this study, effects of capsaicin, an active ingredient of the capsicum plant, were investigated on human 5-hydroxytryptamine type 3 (5-HT₃) receptors. Capsaicin reversibly inhibited serotonin (5-HT)-induced currents recorded by two-electrode voltage clamp method in Xenopus oocytes. The inhibition was time- and concentration-dependent with an IC₅₀ = 62 μM. The effect of capsaicin was not altered in the presence of capsazepine, and by intracellular BAPTA injections or trans-membrane potential changes. In radio-ligand binding studies, capsaicin did not change the specific binding of the 5-HT₃ antagonist [³H]GR65630, indicating that it is a noncompetitive inhibitor of …

Leaf Traits Can Be Used To Predict Rates Of Litter Decomposition, Marc Rosenfield, Jennifer L. Funk, Jason K. Keller, Catrina Clausen, Kimberlee Cyphers

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Strong relationships exist between litter chemistry traits and rates of litter decomposition. However, leaf traits are more commonly found in online trait databases than litter traits and fewer studies have examined how well leaf traits predict litter decomposition rates. Furthermore, while bulk leaf nitrogen (N) content is known to regulate litter decomposition, few studies have explored the importance of N biochemistry fractions, such as protein and amino acid concentration. Here, we decomposed green leaves and naturally senesced leaf litter of nine species representing a wide range of leaf functional traits. We evaluated the ability of traits associated with leaf and …

Hydrogenation Of Organic Matter As A Terminal Electron Sink Sustains High Co2:Ch4 Production Ratios During Anaerobic Decomposition, Rachel M. Wilson, Malak M. Tfaily, Virginia I. Rich, Jason K. Keller, Scott D. Bridgham, Cassandra Medvedeff Zalman, Laura Meredith, Paul J. Hanson, Mark Hines, Laurel Pfeifer-Meister, Scott R. Saleska, Patrick Crill, William T. Cooper, Jeff P. Chanton, Joel E. Kostka

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Once inorganic electron acceptors are depleted, organic matter in anoxic environments decomposes by hydrolysis, fermentation, and methanogenesis, requiring syntrophic interactions between microorganisms to achieve energetic favorability. In this classic anaerobic food chain, methanogenesis represents the terminal electron accepting (TEA) process, ultimately producing equimolar CO₂ and CH₄ for each molecule of organic matter degraded. However, CO₂:CH₄ production in Sphagnum-derived, mineral-poor, cellulosic peat often substantially exceeds this 1:1 ratio, even in the absence of measureable inorganic TEAs. Since the oxidation state of C in both cellulose-derived organic matter and acetate is 0, and CO₂ has …

Enabling Method To Design Versatile Biomaterial Systems From Colloidal Building Blocks, Shalini Saxena, L. Andrew Lyon

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Development of materials with fine spatial control over topographical, mechanical, or chemical features has been investigated for a variety of applications. Here we present a method to fabricate an array of polyelectrolyte constructs including two-dimensionally and three-dimensionally patterned assemblies using both compressible and incompressible colloidal building blocks. This method eliminates prior constraints associated with specific chemistries, and can be used to develop modular, multi-component, patterned assemblies. In particular, development of constructs were investigated using microgels, which are colloidally stable hydrogel microparticles, polystyrene (PS) beads, and PS-microgel core-shell building blocks in conjunction with the polycation poly(ethyleneimine) (PEI). The topography, mechanical properties, …

Temporal Variation In Optical Properties Of Chromophoric Dissolved Organic Matter (Cdom) In Southern California Coastal Waters With Nearshore Kelp And Seagrass, Catherine D. Clark, Warren J. De Bruyn, Paige Aiona

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Optical properties of chromophoric dissolved organic matter (CDOM) were measured in surf zone waters in diurnal field studies at a Southern California beach with nearshore kelp and seagrass beds and intertidal plant wrack. Absorption coefficients (aCDOM(300 nm)) ranged from 0.35 m21 to 3.7 m21 with short-term variability<1 h, increases at ebb and flood tides and higher values (6 m21) during an offshore storm event. Spectral slopes (S) ranged from 0.0028 nm21 to 0.017 nm21, with higher values after the storm; S was generally inversely correlated with aCDOM(300 nm). 3-D excitation–emission matrix spectra (EEMs) for samples with lower S values had humic-type peaks associated with terrestrial material (A, C), marine microbial material (M) and protein peaks, characteristic of freshly produced organic material. Samples with high S values had no or reduced protein peaks, consistent with aged material. Fluorescent indexes (f450/f500 >2.5, BIX>1.1) were consistent with microbial aquatic sources. Leachates of senescent kelp and seagrass had protein and humic-type EEM peaks. After solar simulator irradiation (4 h), protein peaks rapidly photochemically degraded, humic-type peak C increased in intensity and peak M disappeared. Optical characteristics of kelp leachates were most similar to field samples, …

An Upper Limit For Macromolecular Crowding Effects, Andrew C. Miklos, Congang Li, Courtney D. Sorell, L. Andrew Lyon, Gary J. Pielak

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Background: Solutions containing high macromolecule concentrations are predicted to affect a number of protein properties compared to those properties in dilute solution. In cells, these macromolecular crowders have a large range of sizes and can occupy 30% or more of the available volume. We chose to study the stability and ps-ns internal dynamics of a globular protein whose radius is similar to 2 nm when crowded by a synthetic microgel composed of poly(N-isopropylacrylamide-co-acrylic acid) with particle radii of similar to 300 nm.

Results: Our studies revealed no change in protein rotational or ps-ns backbone dynamics and only mild …