Pollen Allergy Suppression Effect by the Oral Administration of Acetic Acid Bacteria (Gluconacetobacter hansenii).

BACKGROUND/AIM: Gluconacetobacter hansenii (G. hansenii) is an acetic acid bacterium of vinegar production. Its anti-allergic effect on mice upon oral administration was examined. MATERIALS AND METHODS: The amount of LPS was measured by the Limulus reaction. Mice were sensitized by peritoneal and intranasal administration of cedar pollen and alum followed by oral administration of 30 or 150 mg/kg of heated G. hansenii cells. Pollen was administered intranasally to evaluate nasal symptoms, and at 8 weeks, IgE and IL-10 levels in blood were measured by ELISA. RESULTS: The amount of LPS in dried bacterial cells was 10.4±3.3 mg/g. In the cedar pollinosis model of mice, a significant reduction was observed in nose scratching of both groups administered with the bacterial cells (30, 150 mg/kg). CONCLUSION: G. hansenii contains LPS, and its oral administration showed an anti-allergic effect by a significant mitigation of the symptoms in a pollen allergy mouse model.

Oral Administration of Lipopolysaccharide of Acetic Acid Bacteria Protects Pollen Allergy in a Murine Model.

BACKGROUND/AIM: Lipopolysaccharide (LPS), a major component of the cell wall of Gram-negative bacteria, is known to possess strong immune-regulatory activity. We have found and reported the existence of biologically-active LPS in acetic acid bacteria. The LPS shows Limulus-positive activity and activation of macrophages to produce nitric oxide and tumor necrosis factor. In this study, we investigated the anti-allergic effect of an orally-administrated acetic acid bacteria extract containing LPS; the cedar pollinosis model was used. MATERIALS AND METHODS: Acetic acid bacteria were isolated from various fruits by Nodai kaihen medium. Then, the anti-allergic effect of acetic acid bacteria extracts was investigated. BALB/c mice were immunized with a mixture of cedar pollen and alum into their peritoneal cavity; they also received additional immunizations of pollen to nasal cavity. After immunizing the mice with pollen into their nasal cavity to trigger an allergic reaction, the frequency of nose scratching was counted for 5 min. RESULTS: The bacteria were cultured and prepared and the water-extract contained about 1-10 mg/ml of Limulus positive substances. The extract of acetic acid bacteria induced higher levels of interleukin (IL)-10 and FOXP3 mRNA expression in macrophages (RAW246.7 cell), as assessed by DNA microarray analysis. Oral administration of the acetic acid bacteria extract demonstrated significantly less scratching numbers than control water group with pollen immunization. CONCLUSION: These results showed that LPS in acetic acid bacteria has the potential to protect from an allergic reaction, especially from cedar pollinosis.

Three-dimensional structure of bovine heart NADH: ubiquinone oxidoreductase (complex I) by electron microscopy of a single negatively stained two-dimensional crystal.

Bovine heart NADH:ubiquinone oxidoreductase (complex I), which is the largest (about 1 MDa) membrane protein complex in the mitochondrial respiratory chain, catalyzes the electron transfer from NADH to ubiquinone, coupled with proton pumping. We have crystallized bovine complex I in reconstituted lipid bilayers and obtained a three-dimensional density map by the electron crystallographic analysis of a single negatively stained two-dimensional crystal. The asymmetric unit with dimensions of a = 388 Å, b = 129 Å and γ = 90° contains two molecules and is of P1 symmetry. Structural differences between the two molecules indicate flexibility of the hydrophilic domain relative to the membrane-embedded domain.

Duplicate polyphenol oxidase genes on barley chromosome 2H and their functional differentiation in the phenol reaction of spikes and grains.

Polyphenol oxidases (PPOs) are copper-containing metalloenzymes encoded in the nucleus and transported into the plastids. Reportedly, PPOs cause time-dependent discoloration (browning) of end-products of wheat and barley, which impairs their appearance quality. For this study, two barley PPO homologues were amplified using PCR with a primer pair designed in the copper binding domains of the wheat PPO genes. The full-lengths of the respective PPO genes were cloned using a BAC library, inverse-PCR, and 3'-RACE. Linkage analysis showed that the polymorphisms in PPO1 and PPO2 co-segregated with the phenol reaction phenotype of awns. Subsequent RT-PCR experiments showed that PPO1 was expressed in hulls and awns, and that PPO2 was expressed in the caryopses. Allelic variation of PPO1 and PPO2 was analysed in 51 barley accessions with the negative phenol reaction of awns. In PPO1, amino acid substitutions of five types affecting functionally important motif(s) or C-terminal region(s) were identified in 40 of the 51 accessions tested. In PPO2, only one mutant allele with a precocious stop codon resulting from an 8 bp insertion in the first exon was found in three of the 51 accessions tested. These observations demonstrate that PPO1 is the major determinant controlling the phenol reaction of awns. Comparisons of PPO1 single mutants and the PPO1PPO2 double mutant indicate that PPO2 controls the phenol reaction in the crease on the ventral side of caryopses. An insertion of a hAT-family transposon in the promoter region of PPO2 may be responsible for different expression patterns of the duplicate PPO genes in barley.

Barley grain with adhering hulls is controlled by an ERF family transcription factor gene regulating a lipid biosynthesis pathway.

In contrast to other cereals, typical barley cultivars have caryopses with adhering hulls at maturity, known as covered (hulled) barley. However, a few barley cultivars are a free-threshing variant called naked (hulless) barley. The covered/naked caryopsis is controlled by a single locus (nud) on chromosome arm 7HL. On the basis of positional cloning, we concluded that an ethylene response factor (ERF) family transcription factor gene controls the covered/naked caryopsis phenotype. This conclusion was validated by (i) fixation of the 17-kb deletion harboring the ERF gene among all 100 naked cultivars studied; (ii) two x-ray-induced nud alleles with a DNA lesion at a different site, each affecting the putative functional motif; and (iii) gene expression strictly localized to the testa. Available results indicate the monophyletic origin of naked barley. The Nud gene has homology to the Arabidopsis WIN1/SHN1 transcription factor gene, whose deduced function is control of a lipid biosynthesis pathway. Staining with a lipophilic dye (Sudan black B) detected a lipid layer on the pericarp epidermis only in covered barley. We infer that, in covered barley, the contact of the caryopsis surface, overlaid with lipids to the inner side of the hull, generates organ adhesion.

Bacillus circulans MH-K1 chitosanase: amino acid residues responsible for substrate binding.

To identify the amino acids responsible for the substrate binding of chitosanase from Bacillus circulans MH-K1 (MH-K1 chitosanase), Tyr148 and Lys218 of the chitosanase were mutated to serine and proline, respectively, and the mutated chitosanases were characterized. The enzymatic activities of Y148S and K218P were found to be 12.5% and 0.16% of the wild type, respectively. When the (GlcN)3 binding ability to the chitosanase was evaluated by fluorescence spectroscopy and thermal unfolding experiments, the binding abilities of both mutant enzymes were markedly reduced as compared with the wild type enzyme. The affinity of the enzyme for the trisaccharide decreased by 1.0 kcal/mol of binding free energy for Y148S, and 3.7 kcal/mol for K218P. The crystal structure of K218P revealed that Pro218 forms a cis-peptide bond and that the state of the flexible loop containing the 218th residue is considerably affected by the mutation. Thus, we conclude that the flexible loop containing Lys218 plays an important role in substrate binding, and that the role of Tyr148 is less critical, but still important, due to a stacking interaction or hydrogen bond.