Resonance Raman study on photoreduction of cytochrome c oxidase: distinction of cytochromes a and a3 in the intermediate oxidation states

Occurrence of photoreduction of bovine cytochrome c oxidase was confirmed with the difference absorption spectra and oxygen consumption measurements for the enzyme irradiated with laser light at 406.7, 441.6, and 590 nm. The resonance Raman spectra were obtained under the same experimental conditions as those adopted for the measurements of oxygen consumption and difference absorption spectra. The photoreduction was more effective upon irradiation at shorter wavelengths and was irreversible under anaerobic conditions. However, upon aeration into the cell, the original oxidized form was restored. It was found that aerobic laser irradiation produces a photo steady state of the catalytic dioxygen reduction and that the Raman scattering from this photo steady state probes cytochrome a2+ and cytochrome a3(3)+ separately upon excitations at 441.6 and 406.7 nm, respectively. The enzyme was apparently protected from the photoreduction in the spinning cell with the spinning speed between 1 and 1500 rpm. These results were explained satisfactorily with the reported rate constant for the electron transfer from cytochrome a to cytochrome a3 (0.58 s-1) and a comparable photoreduction rate of cytochrome a. The anaerobic photoreduction did give Raman lines at 1666 and 214 cm-1, which are characteristic of the ferrous high-spin cytochrome a3(2)+, but they were absent under aerobic photoreduction. The formyl CH = O stretching mode of the a3 heme was observed at 1671 cm-1 for a2+a3(2)+CO but at 1664 cm-1 for a2+a3(2)+CN-, indicating that the CH = O stretching frequency reflects the pi back-donation to the axial ligand similar to the oxidation state marker line (v4).     

Variation in the ribosomal internal transcribed spacers and 5.8S rDNA among five species of Acropora (Cnidaria; Scleractinia): patterns of variation consistent with reticulate evolution

The ITS sequences of Acropora spp. are the shortest so far identified in any metazoan and are among the shortest seen in eukaryotes; ITS1 was 70-80 bases, and ITS2 was 100-112 bases. The ITS sequences were also highly variable, but base composition and secondary structure prediction indicate that divergent sequence variants are unlikely to be pseudogenes. The pattern of variation was unusual in several other respects: (1) two distinct ITS2 types were detected in both A. hyacinthus and A. cytherea, species known to hybridize in vitro with high success rates, and a putative intermediate ITS2 form was also detected in A. cytherea; (2) A. valida was found to contain highly (29%) diverged ITS1 variants; and (3) A. longicyathus contained two distinct 5.8S rDNA types. These data are consistent with a reticulate evolutionary history for the genus Acropora.      

Structural Change of DNA Induced by Nucleoid Proteins: Growth Phase-Specific Fis and Stationary Phase-Specific Dps

The effects of nucleoid proteins Fis and Dps of Escherichia coli on the higher order structure of a giant DNA were studied, in which Fis and Dps are known to be expressed mainly in the exponential growth phase and stationary phase, respectively. Fis causes loose shrinking of the higher order structure of a genome-sized DNA, T4 DNA (166 kbp), in a cooperative manner, that is, the DNA conformational transition proceeds through the appearance of a bimodal size distribution or the coexistence of elongated coil and shrunken globular states. The effective volume of the loosely shrunken state induced by Fis is 30–60 times larger than that of the compact state induced by spermidine, suggesting that cellular enzymes can access for DNA with the shrunken state but cannot for the compact state. Interestingly, Dps tends to inhibit the Fis-induced shrinkage of DNA, but promotes DNA compaction in the presence of spermidine. These characteristic effects of nucleotide proteins on a giant DNA are discussed by adopting a simple theoretical model with a mean-field approximation.     

Double Infection,Recombination, and Segregation of Two R Factors,R 100 and Rts1, and Their Possible Bearing on the Genetic Structure of R 100

Applying the observation by Yokota et al (1969) that a cell doubly harboring an R factor (R100) and a temperature sensitive R factor (Rts1) produces segregant R factors with various resistance patterns, a total of 271 segregant R factors were obtained. There were 163 resistant to (sul, str, kan), 39 resistant to (sul, str, cml, kan), 62 resistant to (sul, str, tet, kan) and finally 7 resistant to (tet, kan). More than 90% of the former 3 segregants were fi⁺ and the remainder, including all of the (tet, kan) segregants, were fi^?. Some fi^? segregants with the former 3 resistance patterns and all of the (tet, kan) segregants were nontransmissible. All of these segregants were still temperature sensitive. Based upon the results of three experiments; (a) the growth at 43 C to observe linked loss of the kan gene and the genes derived from R 100, (b) a conjugal analysis of the relevant resistant markers, and (c) a transductional analysis of these same markers, several conclusions were made. The 2 R factors both consisting of a circle were supposed to have recombined to form a larger circle which then further resulted in the final formation of smaller circles. The possible bearing of these observations and conclusions on the genetic structure of R 100 was discussed.     

Microflow system promotes acetaminophen crystal nucleation

It is known that interfaces have various impacts on crystallization from a solution. Here, we describe crystallization of acetaminophen using a microflow channel, in which two liquids meet and form a liquid–liquid interface due to laminar flow, resulting in uniform mixing of solvents on the molecular scale. In the anti‐solvent method, the microflow mixing promoted the crystallization more than bulk mixing. Furthermore, increased flow rate encouraged crystal formation, and a metastable form appeared under a certain flow condition. This means that interface management by the microchannel could be a beneficial tool for crystallization and polymorph control.     

New clade of silicified bolidophytes that belong to Triparma (Bolidophyceae,Stramenopiles)

The small phytoplankton genus Triparma belongs to the class Bolidophyceae and contains two distinct forms: silicified species and naked flagellated species (formerly Bolidomonas). Recent studies showed that four silicified species/strains (Triparma laevis f. inornata, T. laevis f. longispina, T. strigata, and T. aff. verrucosa) belong to a single clade that is paraphyletic, because it also contains an unclassified flagellated strain, and is sister to a flagellated species, T. eleuthera. In this study, we isolated and characterized two new strains of silicified species to test the phylogenetic unity of silicified bolidophytes. The isolates were identified as T. retinervis strains because they possessed fine areolation on the cell wall. 18S rDNA and rbcL phylogenetic analyses demonstrated that T. retinervis formed a new silicified clade that is sister to the flagellated species T. pacifica. This reveals that there are at least two distinct clades including both silicified and flagellated Triparma species.     

Novel (p)ppGpp0 suppressor mutations reveal an unexpected link between methionine catabolism and GTP synthesis in Bacillus subtilis

In bacteria, guanosine (penta)tetra-phosphate ([p]ppGpp) is essential for controlling intracellular metabolism that is needed to adapt to environmental changes, such as amino acid starvation. The (p)ppGpp⁰ strain of Bacillus subtilis, which lacks (p)ppGpp synthetase, is unable to form colonies on minimal medium. Here, we found suppressor mutations in the (p)ppGpp⁰ strain, in the purine nucleotide biosynthesis genes, prs, purF and rpoB/C, which encode RNA polymerase core enzymes. In comparing our work with prior studies of ppGpp⁰ suppressors, we discovered that methionine addition masks the suppression on minimal medium, especially of rpoB/C mutations. Furthermore, methionine addition increases intracellular GTP in rpoB suppressor and this effect is decreased by inhibiting GTP biosynthesis, indicating that methionine addition activated GTP biosynthesis and inhibited growth under amino acid starvation conditions in (p)ppGpp⁰ backgrounds. Furthermore, we propose that the increase in intracellular GTP levels induced by methionine is due to methionine derivatives that increase the activity of the de novo GTP biosynthesis enzyme, GuaB. Our study sheds light on the potential relationship between GTP homeostasis and methionine metabolism, which may be the key to adapting to environmental changes.     

Life cycle environmental and economic impact of a food waste recycling-farming system: a case study of organic vegetable farming in Japan

Purpose

Bio-based recycling systems and agricultural production using recycled materials are often evaluated separately. This study performs an environmental and socio-economic life cycle assessment (LCA) of a food waste treatment and spinach farming system in Japan. The environmental and economic tradeoffs of introducing a recycling system and the net environmental benefit of the substitution of market fertilizer considering operation changes are also examined.

Methods

Three scenarios were developed and compared. In the conventional (CV) scenario, food waste is collected, incinerated, and disposed of in landfill, and the farmer uses market organic fertilizer. The on-site composting (OC) scenario processes food waste using an on-site garbage disposer and transports compost to a nearby spinach farmer. Food waste in the centralized composting (CC) scenario is transported to a centralized composting facility and resultant compost is sent to the farm. Primary data were obtained from field experiments and interviews. Non-greenhouse gas (GHG) emissions from the field and nitrogen leaching to water systems were simulated using the denitrification–decomposition (DNDC) model.

The environmental LCA targeted climate change, eutrophication, and waste landfill. An input–output analysis estimated socio-economic indicators, namely gross added value and employment inducement effect.

Results and discussion

The scenario with the lowest impact is the CC scenario. Climate change and eutrophication impacts are highest in the OC scenario and waste landfill impacts are most significant in the CV scenario. The weighted impact by LIME2 can be reduced by 47% in the CC scenario and 17% in the OC scenario due to the recycling of food waste instead of dumping in the landfill. The difference in socio-economic indicators between the scenarios was relatively small, although the CV scenario encouraged more employment. The substitution effect of composting, as well as the environmental impact reduction of replacing market organic fertilizer with compost, will result in 28.7% of the avoided impacts in GHG emissions.

Conclusions

Both composting scenarios are feasible from an environmental and socio-economic perspective when compared with conventional organic production, although there is a tradeoff between waste landfill and GHG emissions for the on-site composting system. However, the OC scenario needs to save electricity to improve its environmental competitiveness with the CV scenario. When considering the substitution effect of composting, it is recommended to take into account that agricultural operation also changes.

     

Synthesis and Binding Properties of Oligonucleotides Containing an Azobenzene Linker

Abstract

Incorporation of an azobenzene-4,4′-diamide group via a linker arm into the 3′-hydroxyl function of one oligonucleotide segment and the 5′-OH of other oligonucleotide has been described. The binding of the oligonucleotides containing the azobenzene linker was investigated by UV melting behaviors. The azobenzene linker has been shown to be useful as an effective bridge for stabilizing hairpin duplex and triplex.