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PubMed2004

Crystallization and preliminary X-ray analysis of heparinase II from Pedobacter heparinus.

Heparinase II from Pedobacter heparinus (formerly Flavobacterium heparinum), which acts on both heparin and heparan sulfate, is one of several glycosaminoglycan-degrading enzymes produced by this organism. This enzyme, with a molecular weight of 84 kDa, utilizes a lytic mechanism to cleave the alpha(1-4) glycosidic bond between hexosamine (D-glucosamine) and L-iduronic or D-glucuronic acid, resulting in a product with an unsaturated sugar ring at the non-reducing end. The enzyme was crystallized by the hanging-drop vapour-diffusion method. The crystals belong to orthorhombic space group P2(1)2(1)2(1) and diffract to 2 A resolution. There are two molecules in the asymmetric unit, consistent with the finding that recombinant heparinase II functions as a dimer in solution.

Cloning, Molecular↗

PubMed1978

Humidifier fever and endotoxin exposure.

Three cases of humidifier fever were detected in an office environment. Flavobacteria were found in the contaminated water in a humidifier. After an experimental exposure, the three persons with previous symptoms suffered from fever and slight respiratory symptoms. A leucocytosis and an increase in the number of segmented white blood cells were found the day following the exposure. General immunoglobulins as well as antibodies to Flavobacterium and endotoxin were slightly elevated in the exposed group. The possibility that endotoxins may be the causative agent by means of an indirect complement activation is discussed.

Adult↗

PubMed1986

Isolation and characterization of aerobic heterotrophic bacteria from natural spring waters in the Lanjaron area (Spain).

Aerobic, heterotrophic bacteria were isolated from nine natural mineral water springs in the Lanjaron area of Spain over the period July 1980 to May 1981. The mineral waters contained few bacteria (mean counts 26-5275 cfu per 100 ml) and the bacterial flora of all nine springs was very similar. Most of the isolates were Gram-negative rods (90%), and among these Pseudomonas spp. and members of the Flavobacterium-Cytophaga-Flexibacter group were numerically dominant. Aeromonas-Vibrio and Enterobacteriaceae isolates were an important fraction of the total number, but isolates from remaining groups (Acinetobacter, Chromobacterium, Alcaligenes and Gram-positive organisms) constituted only a small proportion of the flora. The comparatively small number of species isolated, and the occurrence of no more than three or four different bacterial types in spring water of different chemical and physical composition is discussed.

Bacteria, Aerobic↗

PubMed1986

A note on starch hydrolysis and beta-glucuronidase activity among flavobacteria.

Most flavobacteria tested with the fluorogenic substrate 4-methylumbelliferyl-beta-D-glucuronide possessed beta-glucuronidase (GUD), but when some of the same strains were tested with the API ZYM gallery, all were negative for GUD. Conflicting reports also appear in the literature about starch hydrolysis among flavobacteria. We observed that the results obtained can depend on the medium used and the length of incubation. Our results indicate that GUD activity and starch hydrolysis are more widely distributed in the genus Flavobacterium than previously reported.

Culture Media↗

PubMed1989

A numerical taxonomic study of proteolytic and lipolytic psychrotrophs isolated from caprine milk.

Lipolytic and proteolytic psychrotrophs were isolated from raw and pasteurized goats' milk, which had been stored at 5 degrees C for 7 d. The 241 strains isolated and 20 reference strains were examined by 149 biochemical, physiological, and morphological tests. The results yielded 195 characters suitable for taxonomic analysis. Computer-assisted complete linkage analysis, using the Jaccard coefficient, produced 22 phenons at 75% S. The results showed that Pseudomonas fluorescens was the predominant psychrotrophic bacterium, but that Pseudomonas fragi was dominant in some milk samples. Strains of Serratia liquefaciens and Flavobacterium balustinum were also identified.

Animals↗

PubMed1989

Determination of the active-site serine of 6-aminohexanoate-dimer hydrolase.

Diisopropylfluorophosphate, an inhibitor of serine proteinase, was used to label 6-aminohexanoate-dimer hydrolase, a nylon oligomer degradative enzyme of Flavobacterium sp. K172. More than 95% of the enzyme activity was lost upon incorporation of 1-1.5 molecules inhibitor/subunit of the enzyme. The tryptic peptide of the labeled enzyme was purified by HPLC (reverse-phase partition) and its amino acid sequence was identified. Radioactivity was found to be incorporated into an 8-amino-acid peptide (108His-Leu-Leu-Met-Ser-Val-Ser-Lys115). Amino acid alteration from Ser to Ala at the position 112 by site-directed mutagenesis caused loss of enzyme activity to below the detection threshold (1% of the activity of the parental enzyme). These results indicate that Ser112 is essential for the activity.

Affinity Labels↗

PubMed1991

Peptide-N4-(N-acetyl-beta-glucosaminyl)asparagine amidase F cannot release glycans with fucose attached alpha 1----3 to the asparagine-linked N-acetylglucosamine residue.

The ability of peptide-N4-(N-acetyl-beta-glucosaminyl)asparagine amidase F (PNGase F) from Flavobacterium meningosepticum and PNGase A from sweet almonds to deglycosylate N-glycopeptides and N-glycoproteins from plants was compared. Bromelain glycopeptide and horseradish peroxidase-C glycoprotein, which contain xylose linked beta 1----2 to beta-mannose and fucose linked alpha 1----3 to the innermost N-acetylglucosamine, were used as substrates. In contrast to PNGase A, the enzyme from F. meningosepticum did not act upon these substrates even at concentrations 100-fold higher than required for complete deglycosylation of commonly used standard substrates. After removal of alpha 1----3-linked fucose from the plant glycopeptide and glycoprotein by mild acid hydrolysis, they were readily degraded by PNGase F at moderate enzyme concentrations. Hence we conclude that alpha 1----3 fucosylation of the inner N-acetylglucosamine impedes the enzymatic action of PNGase F. Knowledge of this limitation of the deglycosylation potential of PNGase F may turn it from a pitfall into a useful experimental tool.

Acetylglucosamine↗

PubMed1996

Specificity studies of bacterial sulfatases by means of structurally defined sulfated oligosaccharides isolated from shark cartilage chondroitin sulfate D.

Chondro-4-sulfatase and chondro-6-sulfatase from Proteus vulgaris and delta-hexuronate-2-sulfatase from Flavobacterium heparinum are potentially useful tools for structural studies of chondroitin sulfate and dermatan sulfate. Their substrate specificities were investigated with various structurally defined, sulfated hexasaccharides isolated from chondroitin sulfate as described in the accompanying report [Sugahara, K., Nadanaka, S., Takeda, K. & Kojima, T. (1996) Eur. J. Biochem. 239, 871-880]. The results indicated that delta-hexuronate-2-sulfatase released an ester sulfate from the C2 position of the delta-hexuronate residue located at the non-reducing terminus, while chondro-6-sulfatase removed an ester sulfate from the C6 position of the GalNAc residue at the reducing end of the hexasaccharides. Chondro-4-sulfatase acted preferentially on an ester sulfate on the C4 position of the GalNAc residue at the reducing end under mild incubation conditions, but also released a sulfate group under harsh conditions from the C4 position of the GalNAc residue at the internal positions of the hexasaccharide chains, unless the GalNAc residue had another ester sulfate on its C6 position. The results demonstrated the usefulness of the sulfatases as tools for the structural characterization of chondroitin sulfate oligosaccharides.

Animals↗

PubMed2006

Ice-active characteristics of soil bacteria selected by ice-affinity.

As an initial screen for microorganisms that produce ice-active macromolecules, ice-affinity was used to select microorganisms from soil consortia originating from three temperate regions. Once selected and subsequently purified to single colonies, these microbes were putatively identified by 16S ribosomal RNA sequencing and assayed for various ice-active properties. Ice-affinity selection appeared to select for bacteria with ice-associating activities: inhibition of ice recrystallization; ice nucleation; ice shaping. Although none of these activities were observed in Paenibacillus amyloliticus C8, others such as Chryseobacterium sp. GL8, demonstrated both ice recrystallization inhibition and ice-shaping activities. Pseudomonas borealis DL7 was classified as a type I ice nucleator, Flavobacterium sp. GL7, was identified as a type III ice nucleator and Acinetobacter radioresistens DL5 demonstrated ice recrystallization inhibition. In all, 19 different culturable bacteria were selected from the thousands of microbes in late-summer collected soil samples. Many of the selected microbes have been previously reported in glacial ice cores or polar sea ice, and of five isolates that were further characterized, four showed ice-associating activities. These results indicate the significant potential of ice-affinity selection even with temperate climate soils, suggesting that sampling in more extreme and remote areas is not required for the isolation of ice-active bacteria.

Acinetobacter↗

PubMed2007

Isolation and characterization of hydrogen-oxidizing bacteria induced following exposure of soil to hydrogen gas and their impact on plant growth.

In many legumes, the nitrogen fixing root nodules produce H2 gas that diffuses into soil. It has been demonstrated that such exposure of soil to H2 can promote plant growth. To assess whether this may be due to H2-oxidizing microorganisms, bacteria were isolated from soil treated with H2 under laboratory conditions and from soils collected adjacent to H2 producing soybean nodules. Nineteen isolates of H2-oxidizing bacteria were obtained and all exhibited a half-saturation coefficient (Ks) for H2 of about 1 ml l(-1). The isolates were identified as Variovorax paradoxus, Flavobacterium johnsoniae and Burkholderia spp. using conventional microbiological tests and 16S rRNA gene sequence analysis. Seventeen of the isolates enhanced (57-254%) root elongation of spring wheat seedlings. Using an Arabidopsis thaliana bioassay, plant biomass was increased by 11-27% when inoculated by one of four isolates of V. paradoxus or one isolate of Burkholderia that were selected for evaluation. The isolates of V. paradoxus found in both H2-treated soil and in soil adjacent to soybean nodules had the greatest impact on plant growth. The results are consistent with the hypothesis that H2-oxidizing bacteria in soils have plant growth promoting properties.

Arabidopsis↗

PubMed2006

Contribution of ethylamine degrading bacteria to atrazine degradation in soils.

Bacterial communities that cooperatively degrade atrazine commonly consist of diverse species in which the genes for atrazine dechlorination and dealkylation are variously distributed among different species. Normally, the first step in degradation of atrazine involves dechlorination mediated by atzA, followed by stepwise dealkylation to yield either N-ethylammelide or N-isopropylammelide. As the liberated alkylamine moieties are constituents of many organic molecules other than atrazine, it is possible that a large number of alkylamine-degrading bacteria other than those previously described might contribute to this key step in atrazine degradation. To examine this hypothesis, we isolated 82 bacterial strains from soil by plating soil water extracts on agar media with ethylamine as a sole carbon source. Among the relatively large number of isolates, only 3 were able to degrade N-ethylammelide, and in each case were shown to carry the atzB gene and atzC genes. The isolates, identified as Rhizobium leguminosarum, Flavobacterium sp., and Arthrobacter sp., were all readily substituted into an atrazine-degrading consortium to carry out N-ethylammelide degradation. The distribution of these genes among many different species in the soil microbial population suggests that these genes are highly mobile and over time may lead to generation of various atrazine-degrading consortia.

Amidohydrolases↗

PubMed2001

Bacterial community analysis of Indonesian hot springs.

We report the first attempts to describe thermophilic bacterial communities in Indonesia's thermal springs using molecular phylogenetic analyses. 16S rRNA genes from laboratory cultures and DNA directly amplified from three hot springs in West Java were sequenced. The 22 sequences obtained were assignable to the taxa Proteobacteria, Bacillus and Flavobacterium, including a number of clades not normally associated with thermophily.

Bacillus↗

PubMed1978

Antimicrobial susceptibility of flavobacteria.

Antimicrobial susceptibility patterns of 28 clinical isolates of Flavobacterium sp. were determined by standard disk diffusion technique and by antimicrobial dilution in agar. Rifampin, clindamycin, trimethoprim-sulfamethoxazole, cefoxitin, and vancomycin are among the antimicrobial agents which may be clinically useful to treat infections caused by flavobacteria. All 28 isolates were resistant to erythromycin with minimal inhibitory concentrations of 32 mug/ml or more. Currently recommended interpretive zones of inhibition by disk diffusion did not reliably predict antimicrobial susceptibility of the 28 flavobacteria isolates when compared with the agar dilution technique, and, therefore, a more direct measurement of minimal inhibitory or bactericidal concentration is recommended.

Anti-Bacterial Agents↗

PubMed2002

EBR-1, a novel Ambler subclass B1 beta-lactamase from Empedobacter brevis.

Empedobacter brevis (formerly designated Flavobacterium breve) is a gram-negative aerobe involved in nosocomial infections. The Ambler class B beta-lactamase gene bla(EBR-1) was cloned and expressed in Escherichia coli from E. brevis clinical strain ASS-1, which had reduced susceptibility to expanded-spectrum cephalosporins and carbapenems. Purified beta-lactamase EBR-1 hydrolyzed penicillins, cephalosporins, and carbapenems efficiently but not aztreonam. Kinetic parameters of EBR-1 were similar to those of class B enzymes such as BlaB, IND-2, and GOB-1 identified from other Flavobacteriaceae species, except for meropenem, which was more hydrolyzed by beta-lactamase GOB-1. EBR-1, with a pI of 8.0 and a relative molecular mass of ca. 25 kDa, was classified in functional subgroup 3a, which includes most of the class B beta-lactamases. EBR-1, which belongs to molecular subclass B1 of metalloenzymes, shares 58, 57, and 42% amino acid identity with the most closely related beta-lactamases, IND-1/IND-2 from Chryseobacterium indologenes, CGB-1 from Chryseobacterium gleum, and BlaB from Chryseobacterium meningosepticum, respectively.

Amino Acid Sequence↗

PubMed1978

Structural factors influencing the biodegradation of imides.

Comparative studies on the biodegradability of amides and imides are presented. Low-molecular-weight imides of varying chain lengths (4, 6, 7, 8, 18, and 20 carbons) were biodegrable. N-alkyl substitution of amides and imides resulted in non-biodegrable derivatives when the amide portion was greater than two carbons in length. N-alkyl-substituted derivatives of acetamide or diacetamide, however, were biodegrable. Several soil isolates, including Aspergillus niger and species of Flavobacterium and Alcaligenes, were capable of growth with imides as sole N or C sources.

Alcaligenes↗

PubMed1978

Bacterial oxidation of polyethylene glycol.

The metabolism of polyethylene glycol (PEG) was investigated with a synergistic, mixed culture of Flavobacterium and Pseudomonas species, which are individually unable to utilize PEGs. The PEG dehydrogenase linked with 2,6-dichlorophenolindophenol was found in the particulate fraction of sonic extracts and catalyzed the formation of a 2,4-dinitrophenylhydrazine-positive compound, possibly an an aldehyde. The enzyme has a wide substrate specificity towards PEGs: from diethylene glycol to PEG 20,000 Km values for tetraethylene glycol (TEG), PEG 400, and PEG 6,000 were 11, 1.7, and 15 mM, respectively. The metabolic products formed from TEG by intact cells were isolated and identified by combined gas chromatography-mass spectrometry as triethylene glycol and TEG-monocarboxylic acid plus small amounts of TEG-dicarboxylic acid, diethylene glycol, and ethylene glycol. From these enzymatic and analytical data, the following metabolic pathway was proposed for PEG: HO(CH2CH2O)nCH2CH2OH leads to HO(CH2CH2O)nCH2CHO leads to HO(CH2CH2O)nCH2COOH leads to HO(CH2CH2O)n-1CH2CH2OH.

Chemical Phenomena↗

PubMed1983

Bacteriophages of methanotrophs isolated from fish.

Bacteriophages of methanotrophic bacteria were isolated from 67 fish. Only two phages isolated from two fish species specifically lysed Methylocystis sp. and Flavobacterium gasotypicum. The phages lysing these species were designated 63-F and CMF-1-F, respectively. The isolated phages differed greatly in the fine structure of the virion, plaque morphology, spectrum of lytic action, serological properties, and UV sensitivity. At the same time, they had identical one-step growth characteristics: their latent period equalled 5 h, lysis time was 3 to 4 h, and burst size was about 240 virions. The phages had guanine- and cytosine-rich double-stranded DNAs consisting of common nitrogen bases. The molecular masses of the DNAs as determined by the sums of restriction endonuclease cleavage fragments were 28 X 10(6) daltons for phage 63-F and 31 X 10(6) daltons for phage CMF-1-F.

Animals↗

PubMed1984

Purification and partial characterization of Flavotoxin A.

A heat-resistant, low-molecular-weight toxin was isolated from semisolid potato dextrose agar medium after inoculation with Flavobacterium farinofermentans sp. nov., which was isolated from fermented corn meal that caused some outbreaks of food poisoning in China. The toxin was purified by solvent partition, Sephadex LH-20 gel filtration, and C-18 reversed-phase column chromatography. Thin-layer chromatography and high-pressure liquid chromatographic methods were developed for the identification and analysis of the toxin. The purified toxin exhibited a single spot in thin-layer chromatography and a single peak in high-pressure liquid chromatography and had adsorption maxima at 232 and 267 nm. Mass spectral analysis indicated a molecular weight of 169 with an experimental formula of C9H13O3. The 50% lethal dose of purified toxin in mice (oral) was less than 6.84 mg/kg, but greater than 0.68 mg/kg. Postmortem examination showed that the mice died of some type of neurological and cardiovascular system toxicity. The name Flavotoxin A is being assigned to the toxin.

Animals↗