Synthesis of novel heterobranched beta-cyclodextrins from 4(2)-O-beta-D-galactosyl-maltose and beta-cyclodextrin by the reverse action of pullulanase, and isolation and characterization of the products. (1/137)

From the mixture of 4(2)-O-beta-D-galactosyl-maltose (Gal-G2) and beta-cyclodextrin (betaCD), novel heterobranched betaCDs, (Gal-G2)-betaCD and (Gal-G2)2-betaCDs, were synthesized by the reverse action of debranching enzyme. The optimum conditions for the production of (Gal-G2)2-betaCDs were examined. A mixture of (Gal-G2)2-betaCDs was produced in about 4% yield when Aerobacter aerogenes pullulanase (64 units per 1 g of Gal-G2) was incubated with 1.6 M Gal-G2 and 0.16 M betaCD at 50 degrees C for 4 days. The reaction products, (Gal-G2)2-betaCDs, were separated into three peaks by HPLC analysis on a Hypercarb S column. Their structures were analyzed by fast atom bombardment mass spectroscopy and NMR spectroscopies, and confirmed by comparison of their hydrolyzates by beta-galactosidase with the authentic (G2)2 -betaCDs. The structures of (Gal-G2)-betaCD and three components of (Gal-G2)2-betaCDs were identified as 6-O-(GalG2)-betaCD, 6(1),6(2)-, 6(1),6(3)- and 6(1),6(4)-di-O-(Gal-G2)2-betaCD, respectively.  (+info)

Nucleotide sequence of the chromosomal ampC gene of Enterobacter aerogenes. (2/137)

The AmpC beta-lactamase gene and a small portion of the regulatory ampR sequence of Enterobacter aerogenes 97B were cloned and sequenced. The beta-lactamase had an isoelectric point of 8 and conferred cephalosporin and cephamycin resistance on the host. The sequence of the cloned gene is most closely related to those of the ampC genes of E. cloacae and C. freundii.  (+info)

Alternative pathways for siroheme synthesis in Klebsiella aerogenes. (3/137)

Siroheme, the cofactor for sulfite and nitrite reductases, is formed by methylation, oxidation, and iron insertion into the tetrapyrrole uroporphyrinogen III (Uro-III). The CysG protein performs all three steps of siroheme biosynthesis in the enteric bacteria Escherichia coli and Salmonella enterica. In either taxon, cysG mutants cannot reduce sulfite to sulfide and require a source of sulfide or cysteine for growth. In addition, CysG-mediated methylation of Uro-III is required for de novo synthesis of cobalamin (coenzyme B(12)) in S. enterica. We have determined that cysG mutants of the related enteric bacterium Klebsiella aerogenes have no defect in the reduction of sulfite to sulfide. These data suggest that an alternative enzyme allows for siroheme biosynthesis in CysG-deficient strains of Klebsiella. However, Klebsiella cysG mutants fail to synthesize coenzyme B(12), suggesting that the alternative siroheme biosynthetic pathway proceeds by a different route. Gene cysF, encoding an alternative siroheme synthase homologous to CysG, has been identified by genetic analysis and lies within the cysFDNC operon; the cysF gene is absent from the E. coli and S. enterica genomes. While CysG is coregulated with the siroheme-dependent nitrite reductase, the cysF gene is regulated by sulfur starvation. Models for alternative regulation of the CysF and CysG siroheme synthases in Klebsiella and for the loss of the cysF gene from the ancestor of E. coli and S. enterica are presented.  (+info)

Methionine-to-cysteine recycling in Klebsiella aerogenes. (4/137)

In the enteric bacteria Escherichia coli and Salmonella enterica, sulfate is reduced to sulfide and assimilated into the amino acid cysteine; in turn, cysteine provides the sulfur atom for other sulfur-bearing molecules in the cell, including methionine. These organisms cannot use methionine as a sole source of sulfur. Here we report that this constraint is not shared by many other enteric bacteria, which can use either cysteine or methionine as the sole source of sulfur. The enteric bacterium Klebsiella aerogenes appears to use at least two pathways to allow the reduced sulfur of methionine to be recycled into cysteine. In addition, the ability to recycle methionine on solid media, where cys mutants cannot use methionine as a sulfur source, appears to be different from that in liquid media, where they can. One pathway likely uses a cystathionine intermediate to convert homocysteine to cysteine and is induced under conditions of sulfur starvation, which is likely sensed by low levels of the sulfate reduction intermediate adenosine-5'-phosphosulfate. The CysB regulatory proteins appear to control activation of this pathway. A second pathway may use a methanesulfonate intermediate to convert methionine-derived methanethiol to sulfite. While the transsulfurylation pathway may be directed to recovery of methionine, the methanethiol pathway likely represents a general salvage mechanism for recovery of alkane sulfide and alkane sulfonates. Therefore, the relatively distinct biosyntheses of cysteine and methionine in E. coli and Salmonella appear to be more intertwined in Klebsiella.  (+info)

Synthesis of novel heterobranched beta-cyclodextrins from alpha-D-mannosylmaltotriose and beta-cyclodextrin by the reverse action of pullulanase, and isolation and characterization of the products. (5/137)

Alpha-D-mannosyl-maltotriose (Man-G3) were synthesized from methyl alpha-mannoside and maltotriose by the transfer action of alpha-mannosidase. (Man-G3)-betaCD and (Man-G3)2-betaCD were produced in about 20% and 4% yield, respectively when Aerobacter aerogenes pullulanase (160 units per 1 g of Man-G3) was incubated with the mixture of 1.6 M Man-G3 and 0.16 M betaCD at 50 degrees C for 4 days. The reaction products, (Man-G3)-betaCD were separated to three peaks by HPLC analysis on a YMC-PACK A-323-3 column and (Man-G3)2-betaCD were separated to several peaks by HPLC analysis on a Daisopak ODS column. The major product of (Man-G3)-betaCDs was identified as 6-O-alpha-(6(3)-O-alpha-D-mannosylmaltotriosyl)-betaCD by FAB-MS and NMR spectroscopies. The structures of (Man-G3)2-betaCDs were analyzed by TOF-MS and NMR spectroscopies, and confirmed by comparison of elution profiles of their hydrolyzates by alpha-mannosidase and glucoamylase on a graphitized carbon column with those of the authentic di-glucosyl-betaCDs. The structures of three main components of (Man-G3)2-betaCDs were identified as 6(1),6(2)-, 6(1),6(3)- and 6(1),64-di-O-(63-O-alpha-D-mannosyl-maltotriosyl)-betaCD.  (+info)

National epidemiologic surveys of Enterobacter aerogenes in Belgian hospitals from 1996 to 1998. (6/137)

Two national surveys were conducted to describe the incidence and prevalence of Enterobacter aerogenes in 21 Belgian hospitals in 1996 and 1997 and to characterize the genotypic diversity and the antimicrobial resistance profiles of clinical strains of E. aerogenes isolated from hospitalized patients in Belgium in 1997 and 1998. Twenty-nine hospitals collected 10 isolates of E. aerogenes, which were typed by arbitrarily primed PCR (AP-PCR) using two primers and pulsed-field gel electrophoresis. MICs of 10 antimicrobial agents were determined by the agar dilution method. Beta-lactamases were detected by the double-disk diffusion test and characterized by isoelectric point. The median incidence of E. aerogenes colonization or infection increased from 3.3 per 1,000 admissions in 1996 to 4.2 per 1000 admissions in the first half of 1997 (P < 0.01). E. aerogenes strains (n = 260) clustered in 25 AP-PCR types. Two major types, BE1 and BE2, included 36 and 38% of strains and were found in 21 and 25 hospitals, respectively. The BE1 type was indistinguishable from a previously described epidemic strain in France. Half of the strains produced an extended-spectrum beta-lactamase, either TEM-24 (in 86% of the strains) or TEM-3 (in 14% of the strains). Over 75% of the isolates were resistant to ceftazidime, piperacillin-tazobactam, and ciprofloxacin. Over 90% of the strains were susceptible to cefepime, carbapenems, and aminoglycosides. In conclusion, these data suggest a nationwide dissemination of two epidemic multiresistant E. aerogenes strains in Belgian hospitals. TEM-24 beta-lactamase was frequently harbored by one of these epidemic strains, which appeared to be genotypically related to a TEM-24-producing epidemic strain from France, suggesting international dissemination.  (+info)

Growth inhibition caused by overexpression of the structural gene for glutamate dehydrogenase (gdhA) from Klebsiella aerogenes. (7/137)

Two linked mutations affecting glutamate dehydrogenase (GDH) formation (gdh-1 and rev-2) had been isolated at a locus near the trp cluster in Klebsiella aerogenes. The properties of these two mutations were consistent with those of a locus containing either a regulatory gene or a structural gene. The gdhA gene from K. aerogenes was cloned and sequenced, and an insertion mutation was generated and shown to be linked to trp. A region of gdhA from a strain bearing gdh-1 was sequenced and shown to have a single-base-pair change, confirming that the locus defined by gdh-1 is the structural gene for GDH. Mutants with the same phenotype as rev-2 were isolated, and their sequences showed that the mutations were located in the promoter region of the gdhA gene. The linkage of gdhA to trp in K. aerogenes was explained by postulating an inversion of the genetic map relative to other enteric bacteria. Strains that bore high-copy-number clones of gdhA displayed an auxotrophy that was interpreted as a limitation for alpha-ketoglutarate and consequently for succinyl-coenzyme A (CoA). Three lines of evidence supported this interpretation: high-copy-number clones of the enzymatically inactive gdhA1 allele showed no auxotrophy, repression of GDH expression by the nitrogen assimilation control protein (NAC) relieved the auxotrophy, and addition of compounds that could increase the alpha-ketoglutarate supply or reduce the succinyl-CoA requirement relieved the auxotrophy.  (+info)

Changes in the disintegration properties of some brands of paracetamol tablets inoculated with four bacterial species. (8/137)

Four most common brands of paracetamol (4-aceta-midophenol) tablets were examined for the changes in their disintegration properties after inoculation with Staphylococcus aureus, Bacillus cereus, Klebsiella aerogenes and Pseudomonas aeruginosa and incubating for 5 weeks. The disintegration times varied from one brand to the other, reaching maximum values of 72 min., 82 min., 110 min. and 120 min. for S. aureus, B. cereus, P. aeruginosa and Klebsiella aerogenes, respectively. All brands of paracetamol tablets revealed the presence of cotton wool-like fibrils which were seen to be interwoven within the tablets' matrices and these were believed to have caused the higher disintegration times.  (+info)