Journal of Fermentation and Bioengineering最新文献

An endo-polygalacturonase gene was cloned from a mutant of Saccharomyces cerevisiae that can produce a polygalacturonase, and the nucleotide sequence was identified. The polygalacturonase gene consists of 1086 bp. When the polygalacturonase gene was introduced into a strain of S. cerevisiae that cannot produce a polygalacturonase, the transformant secreted a polygalacturonase identical to that produced by the polygalacturonase-producing mutant.

Clavibacter michiganense was identified as a microorganism that hydrolyzed the glucosyl ester linkages at site 19 of steviol glycosides. An enzyme that catalyzes the hydrolysis was purified from the cell-free extract using streptomycin treatment, ammonium sulfate fractionation, Q Sepharose anion exchange chromatography, Sephacryl S-100 gel filtration, and Ether Toyopearl hydrophobic chromatography. The purified enzyme migrated as a single protein band in polyacrylamide gel electrophoresis in the presence and absence of sodium dodecyl sulfate, and isoelectric focusing. The molecular mass was estimated to be approximately 65 kDa, both by gel filtration and sodium dodecyl sulfate/polyacrylamide gel electrophoresis. An isoelectric point, pI, of 4.6, was obtained using isoelectric focusing. The enzyme was most active at around pH 7.5 and at 45°C, and was stable between pH 6–10 and below 40°C. Both Hg²⁺ and p-chloromercuric benzoate inhibited activity. The enzyme hydrolyzed glucosyl ester linkages at site 19 of rebaudioside A, stevioside, rubusoside, and steviol monoglucosyl ester, although it did not cleave 13-O-linked glucosyl residue of rubusoside and steviol monoside. A transglucosylation product having a cellobiosyl residue at site 19 was formed when rubusoside was used as a glucosyl donor and acceptor. The enzyme hydrolyzed glucosidic linkages in p-nitrophenyl β-glucoside faster than glucosyl ester linkages in the steviol glycosides. It also acted on phenyl β-glucoside and salicin, and faintly on sophorobiose and cellobiose. These results indicate that the enzyme is a novel β-glucosidase that hydrolyzes ester linkages.

Two new anthraquinones are described which were produced by stationary cultures of Fusarium oxysporum isolated from the roots of diseased citrus trees. They are 2-acetyl-3,8-dihydroxy- or 3-acetyl-2,8-dihydroxy-6-methoxy-anthraquinone and 2-(1-hydroxyethyl)-3,8-dihydroxy- or 3-(1-hydroxyethyl)-2,8-dihydroxy-6-methoxy-anthraquinone. These compounds differ from most other microbial anthraquinones in that they lack hydroxyl substituents at the 1,4 positions, and have an acetyl or 1-hydroxyethyl group at the 2 or 3 positions. This is the first report of anthraquinone metabolites produced by F. oxysporum.

A sulfite resistance gene, SSU1-R, was cloned from a highly sulfite-resistant strain of Saccharomyces cerevisiae, Y-9, which was isolated as a contaminant from a wine must. The coding region sequence of SSU1-R was almost identical to that of SSU1, the gene that has been shown to be responsible for sulfite resistance. Although SSU1 is located on chromosome XVI of S. cerevisiae, the upstream region of SSU1-R was homologous with a sequence of chromosome VIII. In our Northern analysis, an intense band was detected in Y-9 and K1-V1116 strains, both of which exhibit strong sulfite resistance, with and without sulfite in the medium. On the other hand, under the same experimental conditions, no band was detected in OC-2, a strain that exhibits weak sulfite resistance. Physical mapping of SSU1/SSU1-R showed that this sequence was on chromosome VIII of Y-9 and the two wine strains, K1-V1116 and WE14. OC-2 and twenty-three other wine, sake, beer, shochu (Japanese distilled liquor), alcohol, bakery, and laboratory strains, had this sequence on chromosome XVI, and four other wine strains had it on both chromosomes. Thus, the difference in the upstream sequence of SSU1/SSU1-R seems to cause differences in the transcription rates and degree of sulfite resistance.

Using four kinds of antibiotics, an investigation was made to determine whether or not H₂O₂ generated from a ZnO powder slurry was related to its antibacterial activity. Changes in the sensitivity of Escherichia coli to the antibiotics suggested that H₂O₂ was one of the primary factors concerned in the antibacterial activity of the ZnO powder slurry.

The performance of photosynthetic electrochemical cells using Anabaena variabilis M-3 immobilized within alginate beads was investigated during repeated discharge and culture cycles of 10-h each. The effect of the light intensity during the culture periods on the duration of the current output and the recovery of endogenous total sugar in the Anabaena cells was examined. Compared with continuous discharge operation, the duration of the current output was extenced by 10 times when the electrochemical cell was operated under 10-h discharge (dark)/culture (light) cycles with a light intensity of 100 W/m² during the culture periods. The conversion efficiency of light energy into electricity was about 0.2% under these conditions.

Acetic acid inhibited the formation of cytochromes and the respiratory activity of the halo-tolerant yeast Zygosaccharomyces rouxii R-1. The growth of the yeast was significantly inhibited by 0.5% acetic acid in a medium containing 18% NaCl. This growth inhibition by acetic acid is thought to be related to the halotolelance of the yeast. Proton expulsive activity, which is necessary for the yeast cells to be able to tolerate and grow in a high-saline environment, was markedly lowered in the presence of 0.5% acetic acid, although the plasma membrane ATPase, which drives proton expulsion, seemed not to be inhibited by 0.5% acetic acid. It is assumed that halo-tolerant yeasts are less tolerant to high concentrations of NaCl in the presence of acetic acid because of the reduction of proton expulsive activity, thereby inhibiting growth.

In spite of its importance, little attention has been paid thus far to the toxicity of chlorinated aliphatic hydrocarbons under methanogenic conditions. The effect of several parameters which can affect the toxicity of CHCl₃, CCl₄, C₂H₂Cl₄ and trans-C₂H₂Cl₂ on the acetoclastic methanogenic activity of anaerobic sludge was studied. The following conclusions were derived: (i) The extend of inhibition produced is a function of the toxicant concentration, the type and the amount of the biomass. A lag-period was detected for the inhibition produced by CHCl₃ and trans-C₂H₂Cl₂, while for the other compounds the inhibitory effect was instant. (ii) The recovery time for an anaerobic granular sludge exposed to different toxicants was a function of the extent of inhibition and the exposure time, and seemed to be independent of the toxicants used. (iii) The possible acclimatization of an anaerobic sludge to continuous exposure to a toxicant was dependent on the nature of the toxicant. The time required to acquire an activity similar to that of untreated controls was dependent on the concentration of the toxicant used. In our case adaptation was only obtained for CCl₄.

A 5.2-kb DNA fragment containing the upstream region of the nitrite reductase gene (nirS) was cloned from Paracoccus denitrificans IFO 12442 and its DNA sequence was determined. In this fragment, four open reading frames (ORFs) were observed. Among these, two ORFs, located 3 kb upstream of the nirS gene were found to encode nitric oxide reductase (norC, norB) by homology analysis. The norC and norB encoded cytochrome c and b subunits of the enzyme, and the predicted molecular weights of the protein products were 17 kDa (150 amino acid residues) and 52.5 kDa (462 amino acid residues), respectively. The other two open reading frames, designated ORF1 (73 kDa, 681 amino acid residues) and ORF2 (32.5 kDa, 304 amino acid residues), were found between the nir and nor genes. The deduced amino acid sequence of ORF1 showed high similarity (35% identity) to that of NosR which is known to be a transcriptional regulatory protein for the N₂O reductase gene cluster of Pseudomonas stutzeri. Since the ORF1 protein has a well conserved cysteine cluster and 5 hydrophobic transmembrane repeats similar to those of the NosR protein, ORF1 probably functions as a membrane bound sensor for denitrification. Northern blot analysis indicated that the ORF1–2 and norCB regions are probably transcribed as independent operons.