Nutrient acquisition and limitation for the photoautotrophic growth of Synechocystis sp. PCC6803 as a renewable biomass source

Photoautotrophic microorganisms (cyanobacteria and algae) offer high promise as a source of biomass for renewable energy due to their rapid growth rates and high biomass yields. To provide a framework for evaluating the feasibility of growing phototrophic microorganisms with high biomass production rates, we operated a bench‐scale photobioreactor using Synechocystis sp. PCC6803 and with light conditions imitating actual day–night light irradiance (LI). During the time of peak LI, PCC6803's specific growth rate (1.7 day⁻¹) and the nitrate uptake rate (0.46 g N/g DW day) were high compared to past reports. Analysis employing the stoichiometry of photosynthesis of PCC6803 and ionic speciation showed that bicarbonate and phosphate were driven to very low concentrations for the high‐LI conditions. In particular, the systematic evaluation of rate‐limiting factors identified when the CO₂–C_i supply rate needed to be increased to mitigate HCO depletion and a large pH increase. It also showed that the traditional BG‐11 medium needs to be augmented with phosphate to avoid severe P depletion. This work exploits quantitative understanding the stoichiometry and kinetics of cyanobacteria for the high‐rate production of a renewable biomass. Biotechnol. Bioeng. 2011;108: 277–285. © 2010 Wiley Periodicals, Inc.     

Conformation of poly(L-arginine). I. Effects of anions

The conformational transition of poly(L -agrignine) by binding with various mono-, di-, and polyvalent anions, especially with SO, was studied by CD measurements. The intramolecular random coil-to-α-helix conformational transition and the subsequent transition to the β-turn-like structure was caused by binding with SO. The binding data obtained from equilibrium dialysis experiments showed that the α-helical conformation of poly(L -arginine) is stabilized at a 1:3 stoichiometric ratio of bound SO to arginine residue; at higher free SO concentrations, the α-helix converts to the β-turn-like structure accompanied by a decrease in amount of bound SO. The same conformaitonal transition of poly(L -arginine) also occurred in the solutions of other divalent anions (SO, CO, and HPO) and polyvalent anions (P₂O, P₃O). Among the monovalent anions examined, CIO and dodecyl sulfate were effective in including α-helical conformation, while the other monovalent anions (OH^?, Cl^?, F^?, H₂PO, HCO and CIO) failed to induce poly(L -arginine) to assume the α-helical conformation. Thus, we noticed that, except for dodecyl sufate, the terahedral structure is common to the α-helix-forming anions. A well-defined model to the α-helical poly(L -arginine)/anion complex was proposed, in which both the binding stoichiometry of anions to the arginine residue and the tetrahedral structure of anions were taken into consideration. Based on these results, it was concluded that the tetrahedral-type anions stabilize the α-helical conformation of poly(L -arginine) by crosslinking between two guanidinium groups of nearby side chains on the same α-helix through the ringed structures stabilized by hydrogen bonds as well as by electrostatic interaction. Throughout the study it was noticed that the structural behavior of poly(L -arginine) toward anions is distinct from that of poly(L -lysine).     

Étude expérimentale de l'auto-association des molécules modèles dipeptidiques. II. Association stéréosélective des molécules énantiomères

Modes of aggregation fo alanine-, norvaline- and valine-contaiing dpepetides with the general formula R₁? C₁O₁? N₂H₂? CHR₂? C₂O₂? N₃H₃? R₃ have been studied in CCl₄ solution by using infrared and nuclear magnetic resonance spectroscopies. Solutions of the pure L isomer and of the racemic mixture do not give identical data. At a given concentration, the racemic mixtrue is more aggregated than the pure enantiomer, and the difference, negligible in the case of alanine derivative, increases wiht the bulkiness of the side cahin R₂. The results show that a selective interaction takes place between enantiomeric molecules, resulting ina dimer associating tow inverse configurated C₅ conformers. The stabilizaion of this dimer proceeds from two symmetrical and intermolecular H₃ … O₁ hydrogen bondings.     

How Large is the Probability for the Estimate of a Variance Component to be Negative?

For a balanced one-way classification, where the normally distributed observations obey a random model y_ij=μ+b_i+c_ij with two variance components var (b_i) = δ and var (c_ij) = δ, the probability is given that the analysis of variance estimate of δ will be negative. This probability depends on δ/δ and the degrees of freedom in the ANOVA table. Tables for this probability are given. If the normally distributed observations obey an intra-class correlation model, the probability that the Mean Square between groups is smaller than the Mean Square within groups can also be evaluated from the given tables.     

Nitrate or ammonium: Influences of nitrogen source on the physiology of a green alga

In freshwaters, algal species are exposed to different inorganic nitrogen (N_i) sources whose incorporation varies in biochemical energy demand. We hypothesized that due to the lesser energy requirement of ammonium ()‐use, in contrast to nitrate ()‐use, more energy remains for other metabolic processes, especially under CO₂‐ and phosphorus (P_i) limiting conditions. Therefore, we tested differences in cell characteristics of the green alga Chlamydomonas acidophila grown on or under covariation of CO₂ and P_i‐supply in order to determine limitations, in a full‐factorial design. As expected, results revealed higher carbon fixation rates for ‐grown cells compared to growth with under low CO₂ conditions. ‐grown cells accumulated more of the nine analyzed amino acids, especially under P_i‐limited conditions, compared to cells provided with . This is probably due to a slower protein synthesis in cells provided with . In contrast to our expectations, compared to ‐grown cells ‐grown cells had higher photosynthetic efficiency under P_i‐limitation. In conclusion, growth on the N_i‐source did not result in a clearly enhanced C_i‐assimilation, as it was highly dependent on P_i and CO₂ conditions (replete or limited). Results are potentially connected to the fact that C. acidophila is able to use only CO₂ as its inorganic carbon (C_i) source.     

On the “filterable aggregates and other particles” interpretation of the extraordinary regime of polyelectrolytes

Quasi-elastic light scattering studies on some polyelectrolyte systems exhibit a somewhat “bizarre” behavior in the profile of the apparent diffusion coefficient D_app as a function of the salt concentration C_s. As C_s is decreased, D_app first increases in accordance with polyelectrolyte theories, and then undergoes a precipitous drop in value by over an order of magnitude at a well-defined critical value C_s = C. This “transition” from C_s > C (ordinary) to C_s < C (extraordinary) is referred to as the “ordinary-extraordinary” (o-e) transition. Ghosh, Peitzsch, and Reed [(1992) Biopolymers, Vol. 32, pp. 1105–1122] proposed a “filterable aggregate” (FA) and “other particle” interpretation for the o-e transition and its reversibility in regard to ionic strength changes. The present communication examines in detail the FA model as applied to the o-e transition. It is shown that the FA model fails to account of the established characteristics of the o-e transition. © 1993 John Wiley & Sons, Inc.     

Phenotypic characterization of transgenic mice harboring Nf1+/- or Nf1-/- osteoclasts in otherwise Nf1+/+ background

Skeletal abnormalities in neurofibromatosis type 1 syndrome (NF1) are observed in ～50% of patients. Here, we describe the phenotype of Nf1_Ocl mouse model with Nf1‐deficient osteoclasts. Nf1_Ocl mice with Nf1^+/? or Nf1^?/? osteoclasts in otherwise Nf1^+/+ background were successfully generated by mating parental Nf1flox/flox and TRAP‐Cre mice. Contrary to our original hypothesis, osteoporotic or fragile bone phenotype was not observed. The µCT analysis revealed that tibial bone marrow cavity, trabecular tissue volume, and the perimeter of cortical bone were smaller in Nf1 mice compared to Nf1 control mice. Nf1 mice also a displayed narrowed growth plate in the proximal tibia. In vitro analysis showed increased bone resorption capacity and cytoskeletal changes including irregular cell shape and abnormal actin ring formation in Nf1^?/? osteoclasts. Surprisingly, the size of spleen in Nf1 mice was two times larger than in controls and histomorphometric analysis showed splenic megakaryocytosis. In summary, Nf1Ocl mouse model presented with a mild but specific bone phenotype. This study shows that NF1‐deficiency in osteoclasts may have a role in the development of NF1‐related skeletal abnormalities, but Nf1‐deficiency in osteoclasts in Nf1^+/+ background is not sufficient to induce skeletal abnormalities analogous to those observed in patients with NF1. J. Cell. Biochem. 113: 2136–2146, 2012. © 2012 Wiley Periodicals, Inc.     

Conformational analysis of acetyl-L-phenylalanine p-acetyl and p-valeryl anilides

Empirical force-field calculations and ir and ¹H-nmr spectra indicate that five-membered (C₅) and seven-membered (C) hydrogen-bonded rings are the preferred conformations of acetyl-L -Phe p-acetyl and p-valeryl anilides in nonpolar media. The C₅/C ratio was found to be dependent on the dryness of the solute and the solvent. This fact and the results from conformational-energy calculations suggest that a molecule of water participates in the stabilization of the C conformation.     

Reactivity of Hg(II) with superoxide: Evidence for the catalytic dismutation of superoxide by HG(II)

Mercuric ion, a well-known nephrotoxin, promotes oxidative tissue damage to kidney cells. One principal toxic action of Hg(II) is the disruption of mitochondrial functions, although the exact significance of this effect with regard to Hg(II) toxicity is poorly understood. In studies of the effects of Hg(II) on superoxide (O) and hydrogen peroxide (H₂O₂) production by rat kidney mitochondria, Hg(II) (1–6 μM), in the presence of antimycin A, caused a concentration-dependent increase (up to fivefold) in mitochondrial H₂O₂ production but an apparent decrease in mitochondrial O production. Hg(II) also inhibited O-dependent cytochrome c reduction (IC₅₀ ≈?2–3 μM) when O was produced from xanthine oxidase. In contrast, Hg(I) did not react with O in either system, suggesting little involvement of Hg(I) in the apparent dismutation of O by Hg(II). Hg(II) also inhibited the reactions of KO₂ (i.e., O) with hemin or horseradish peroxidase dissolved in dimethyl sulfoxide (DMSO). Finally, a combination of Hg(II) and KO₂ in DMSO resulted in a stable UV absorbance spectrum [currently assigned Hg(II)-peroxide] distinct from either Hg(II) or KO₂. These results suggest that Hg(II), despite possessing little redox activity, enhances the rate of O dismutation, leading to increased production of H₂O₂ by renal mitochondria. This property of Hg(II) may contribute to the oxidative tissue-damaging properties of mercury compounds.     

Exploration of the active site of Escherichia coli cystathionine γ‐synthase

Cystathionine γ‐synthase (CGS) catalyzes the condensation of O‐succinyl‐L ‐homoserine (L ‐OSHS) and L ‐cysteine (L ‐Cys), to produce L ‐cystathionine (L ‐Cth) and succinate, in the first step of the bacterial transsulfuration pathway. In the absence of L ‐Cys, the enzyme catalyzes the futile α,γ‐elimination of L ‐OSHS, yielding succinate, α‐ketobutyrate, and ammonia. A series of 16 site‐directed variants of Escherichia coli CGS (eCGS) was constructed to probe the roles of active‐site residues D45, Y46, R48, R49, Y101, R106, N227, E325, S326, and R361. The effects of these substitutions on the catalytic efficiency of the α,γ‐elimination reaction range from a reduction of only ～2‐fold for R49K and the E325A,Q variants to 310‐ and 760‐fold for R361K and R48K, respectively. A similar trend is observed for the k_cat/K of the physiological, α,γ‐replacement reaction. The results of this study suggest that the arginine residues at positions 48, 106 and 361 of eCGS, conserved in bacterial CGS sequences, tether the distal and α‐carboxylate moieties, respectively, of the L ‐OSHS substrate. In contrast, with the exception of the 13‐fold increase observed for R106A, the K is not markedly affected by the site‐directed replacement of the residues investigated. The decrease in k_cat observed for the S326A variant reflects the role of this residue in tethering the side chain of K198, the catalytic base. Although no structures exist of eCGS bound to active‐site ligands, the roles of individual residues is consistent with the structures inhibitor complexes of related enzymes. Substitution of D45, E325, or Y101 enables a minor transamination activity for the substrate L ‐Ala.     

Free‐energy profiles for ions in the influenza M2‐TMD channel

M₂ transmembrane domain channel (M₂‐TMD) permeation properties are studied using molecular dynamics simulations of M₂‐TMD (1NYJ) embedded in a lipid bilayer (DMPC) with 1 mol/kg NaCl or KCl saline solution. This study allows examination of spontaneous cation and anion entry into the selectivity filter. Three titration states of the M₂‐TMD tetramer are modeled for which the four His³⁷ residues, forming the selectivity filter, are net uncharged, +2 charged, or +3 charged. M₂‐TMD structural properties from our simulations are compared with the properties of other models extracted from NMR and X‐ray studies. During 10 ns simulations, chloride ions occasionally occupy the positively‐charged selectivity filter region, and from umbrella sampling simulations, Cl^? has a lower free‐energy barrier in the selectivity‐filter region than either Na⁺ or NH, and NH has a lower free‐energy barrier than Na⁺. For Na⁺ and Cl^?, the free‐energy barriers are less than 5 kcal/mol, suggesting that the 1NYJ conformation would probably not be exquisitely proton selective. We also point out a rotameric configuration of Trp⁴¹ that could fully occlude the channel. Proteins 2009. © 2009 Wiley‐Liss, Inc.     

Heat capacity changes and partial molal heat capacities of some di- and tripeptides in water

Integral enthalpies of solution of several dipeptides and tripeptides in water at low concentrations have been determined at 25 and 35°C. These data have been used to derive the changes in heat capacity on dissolution at infinite dilution ΔC at 30°C. Limiting partial molal heat capacities ΔC have been determined by combining ΔC with C_p2 (heat capacity of pure solid peptides). Using the data on ω-amino acids and these peptides, the partial molal heat capacity of a peptide group ? CONH? was semiquantitatively estimated.     

Einfluß extracellulärer Kaliuminoenkonzentrationen auf die celluläre Natriumionenkonzentration von Lodderomyces elongisporus D

It was found that the cellular Na⁺-concentration (C) of Lodderomyces elongisporus D is depended on the extracellular K⁺-concentration (C). The relationship can be described by an equation in the form The function of the natrium ion seem to be to support the utilisation rate of potassium ion at lower extracellular K⁺-concentration.     

An Unbalanced Mixed Model with Random Effects Having Unequal Variances

Consider the mixed model where x_ijk's are known constants, β_k are unknown parameters and a_i, e_ij are random variables independently and normally distributed with zero means and variances σd_i and σ² respectively, where it is assumed that the d_i's are known (d_i >0). This paper presents procedures for estimating the variance components σ, σ², for testing the hypothesis σ = 0, and for making transformations to random variables with uncorrelated errors and constant variances in order to estimate as well as to test hypothesis concerning the β_k's in the model.     

Effects of temperature on microbial ethanol production. II. A thermodynamic interpretation of the temperature profile curve of ethanol production

An attempt is made to give a thermodynamic interpretation of the complete temperature profile curve of ethanol formation. Taking into consideration an enhancing competition between thermal activation and thermal deactivation of ethanol formation at increasing temperatures and supposing that the ethanol production is affected by a reversible and an irreversible term of thermal deactivation of a modified ARRHENIUS equation being current for the total biokinetic sphere may be derived: . The quantities ΔH and ΔH^D_2T are identical with the temperature functions of the change of entropy caused by reversible and irreversible deactivation of ethanol formation, respectively. Accordingly for the yeast strain Saccharomyces cerevisiae Sc 5 the calculated entropy coefficients of reversible and irreversible thermal deactivation of ethanol formation amount to C = (0.245± 0.013) kJ/mol · deg.² and C = (1.657 ± 0.046) kJ/mol · deg.².     

Characterization of site‐directed mutants of residues R58, R59, D116, W340 and R372 in the active site of E. coli cystathionine β‐lyase

Cystathionine β‐lyase (CBL) catalyzes the hydrolysis of L ‐cystathionine (L ‐Cth) to produce L ‐homocysteine, pyruvate, and ammonia. A series of active‐site mutants of Escherichia coli CBL (eCBL) was constructed to investigate the roles of residues R58, R59, D116, W340, and R372 in catalysis and inhibition by aminoethoxyvinylglycine (AVG). The effects of these mutations on the k_cat/K for the β‐elimination reaction range from a reduction of only 3‐fold for D116A and D116N to 6 orders of magnitude for the R372L and R372A mutants. The order of importance of these residues for the hydrolysis of L ‐Cth is: R372 >> R58 > W340 ≈ R59 > D116. Comparison of the kinetic parameters for L ‐Cth hydrolysis with those for inhibition of eCBL by AVG demonstrates that residue R58 tethers the distal carboxylate group of the substrate and confirms that residues W340 and R372 interact with the α‐carboxylate moiety. The increase in the pK_a of the acidic limb and decrease in the pK_a of the basic limb of the k_cat/K versus pH profiles of the R58K and R58A mutants, respectively, support a role for this residue in modulating the pK_a of an active‐site residue.     

Evaluating four mathematical models for nitrous oxide production by autotrophic ammonia‐oxidizing bacteria

There is increasing evidence showing that ammonia‐oxidizing bacteria (AOB) are major contributors to N₂O emissions from wastewater treatment plants (WWTPs). Although the fundamental metabolic pathways for N₂O production by AOB are now coming to light, the mechanisms responsible for N₂O production by AOB in WWTP are not fully understood. Mathematical modeling provides a means for testing hypotheses related to mechanisms and triggers for N₂O emissions in WWTP, and can then also become a tool to support the development of mitigation strategies. This study examined the ability of four mathematical model structures to describe two distinct mechanisms of N₂O production by AOB. The production mechanisms evaluated are (1) N₂O as the final product of nitrifier denitrification with NO as the terminal electron acceptor and (2) N₂O as a byproduct of incomplete oxidation of hydroxylamine (NH₂OH) to NO. The four models were compared based on their ability to predict N₂O dynamics observed in three mixed culture studies. Short‐term batch experimental data were employed to examine model assumptions related to the effects of (1) NH concentration variations, (2) dissolved oxygen (DO) variations, (3) NO accumulations and (4) NH₂OH as an externally provided substrate. The modeling results demonstrate that all these models can generally describe the NH, NO, and NO data. However, none of these models were able to reproduce all measured N₂O data. The results suggest that both the denitrification and NH₂OH pathways may be involved in N₂O production and could be kinetically linked by a competition for intracellular reducing equivalents. A unified model capturing both mechanisms and their potential interactions needs to be developed with consideration of physiological complexity. Biotechnol. Bioeng. 2013; 110: 153–163. © 2012 Wiley Periodicals, Inc.     

On the blue color of triiodide ions in starch and starch fractions. I. Characterization and assignment of absorption and circular dichroism spectra of triiodide ions in amylose

Four fundamental Raman lines were observed at 159, 111, 55 and 27 cm^-1 corresponding to the I bound (I) in amyloses with DP from 20 to 100, regardless of the degree of polymerization of I and the excitation wavelength. The spectral resolution was based on the molar extinction coefficient and molar ellipticity spectra of I. Eight bands, named, S₁, S₂, ?, S₈ from long to short wavelength, were isolated. These were found regardless of the DP. By a resonance excitation Raman study, the characteristics of S₃ and S₄, comprising the shoulder around 480 nm, were found to be different from those of S₁ and S₂, comprising the blue band. The assignment of the spectra was based on the electronic states of the monomeric I in the exciton-coupled dimeric unit. It was concluded that the blue band (S₁,S₂) belonged to the long-axis transitions and the shoulder band (S₃,S₄) to the short-axis ones on the monmeric coordinate system.     

Apparent molar volumes of some amino acids and peptides in aqueous urea solutions

Densities of solutions of several α-amino acids and peptides in 3 and 6m aqueous urea solvents have been determined at 298.15 K. These data have been used to evaluate the infinite-dilution apparent molar volumes of the solutes and the volume changes due to transfer (V ) of the α-amino acids and peptides at infinite dilution from water to aqueous urea solutions. The sign and magnitude of the V values have been rationalized in the framework of Friedman's cosphere-overlap model. The V values for the glycyl group (? CH₂CONH? ) and alkyl side chains have been estimated.