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PubMed2003

[Purification and characterization of a uridine phosphorylase from Enterobacter aerogenes EAM-Z1].

A uridine phosphorylase(UPase) was isolation from Enterobacter aerogenes EAM-Z1 and purified by means of ammonium sulfate precipitation, DEAE-cellulose, Phenyl-Sepharose, DEAE-Sepharose, FPLC ion exchange, and Sephacryl S-200 column chromatography. The purified UPase showed homogeneity on the native polacrylamide gel electrophoresis. The UPase is a trimer of 43 kD subunits. Fifteen residues from the amino terminal end of UPase were identified as MRMVDLIATKRDGGE. The isoelectric point was pH 4.46. Michaelis constant for uridine was 0.29 mmol/L. The UPase has a maximal activity at a pH value of 7.8 and 50 degrees C. The UPase could catalyses the phosphorolysis of uridine, thymidine, 5-Fluorouridine, 5-Fluoro-2'-deoxyuridine, uracil-beta-D-arbinofuranoside, and could also catalyse the synthesis of 5-Fluorouridine, a better prodrug form of the anticancer drug 5-fluorouracil, from 5-fluorouracil and uridine, and 47% uridine was converted to 5-Fluoro-uridine.

Bacterial Proteins↗
PubMed1992

Purification and characterization of indolepyruvate decarboxylase. A novel enzyme for indole-3-acetic acid biosynthesis in Enterobacter cloacae.

Indolepyruvate decarboxylase, a key enzyme for indole-3-acetic acid biosynthesis, was found in extracts of Enterobacter cloacae. The enzyme catalyzes the decarboxylation of indole-3-pyruvic acid to yield indole-3-acetaldehyde and carbon dioxide. The enzyme was purified to apparent homogeneity from Escherichia coli cells harboring the genetic locus for this enzyme obtained from E. cloacae. The results of gel filtration experiments showed that indolepyruvate decarboxylase is a tetramer with an M(r) of 240,000. In the absence of thiamine pyrophosphate and Mg2+, the active tetramers dissociate into inactive monomers and dimers. However, the addition of thiamine pyrophosphate and Mg2+ to the inactive monomers and dimers results in the formation of active tetramers. These results indicate that the thiamine pyrophosphate-Mg2+ complex functions in the formation of the tetramer, which is the enzymatically active holoenzyme. The enzyme exhibited decarboxylase activity with indole-3-pyruvic acid and pyruvic acid as substrates, but no decarboxylase activity was apparent with L-tryptophan, indole-3-lactic acid, beta-phenylpyruvic acid, oxalic acid, oxaloacetic acid, and acetoacetic acid. The Km values for indole-3-pyruvic acid and pyruvic acid were 15 microM and 2.5 mM, respectively. These results indicate that indole-3-acetic acid biosynthesis in E. cloacae is mediated by indolepyruvate decarboxylase, which has a high specificity and affinity for indole-3-pyruvic acid.

Carboxy-Lyases↗
PubMed1975

Purification and properties of a phosphohydrolase from Enterobacter aerogenes.

A phosphohydrolase from Enterobacter aerogenes which hydrolyzes phosphate mono- and diesters has been purified approximately 50-fold to apparent homoeneity and crystallized. The enzyme is produced when the bacteria utilize phosphate diesters as sole phosphorus source. From sedimentation equilibrium experiments the molecular weight of the native enzyme is 173,000; from sodium dodecyl sulfate polyacrylamide gel electrophoresis the subunit molecular weight is 29,000, indicating that the enzyme is hexameric. The hydrolytic activity of the enzyme using both mono- and diesters is maximal at pH 5; THE Km of the enzyme for bis-p-nitrophenyl phosphate is constant from pH 5 to 8.5 whereas that for p-nitrophenyl phosphate increases about 40-fold as the pH increases over the same range. The phosphodiesterase activity is not inhibited by chelating agents but is inhibited by several divalent metal ions. 31-P NMR spectroscopy was used to identify the hydrolysis products of glycoside cyclic phosphates. The enzyme-catalyzed hydrolysis of methyl beta-D-ribofuranoside cyclic 3:5-phosphate yields exclusively the 5-phosphate whereas that of adenosine 3:5-monophosphate yields a 4:1 mixture of 3- and 5- AMP.

4-Nitrophenylphosphatase↗
PubMed2006

[Pyogenic spondylitis by Enterobacter cloacae as a postoperative complication of TURP: a case report].

We report a case of pyogenic spondylitis caused by Enterobacter cloacae as a rare complication of transurethral resection of the prostate (TURP). A 79-year-old man underwent TURP. Immediate after removal of urethral catheter on postoperative day (POD) 7, he developed high fever > 40 degrees C with increased acute inflammatory reaction. Urine and blood culture detected E. cloacae and methicillin-resistant Staplylococcus aureus. He complained of lumbago since POD 9. Two-week administration of imipenem and teicoplanin resulted in resolution of fever as well as laboratory data, so intravenous antibiotics were changed to oral gatifloxacin. However, his lumbago worsened and gait disturbance appeared. On POD 39, diagnosis of pyogenic spondylitis was finally obtained by Ga-scintigraphy and magnetic resonance imaging. Aspiration of the intervertebral disk (L4-5) revealed E. cloacae as the causative organism of pyogenic spondylitis. His condition improved after conservative treatment with teicoplanin, meropenem and ciplofloxacin for 9 weeks.

Aged↗
PubMed1991

Carbon-phosphorus bond cleavage activity in cell-free extracts of Enterobacter aerogenes ATCC 15038 and Pseudomonas sp. 4ASW.

Carbon-phosphorus bond cleavage activity was investigated in cell-free extracts of Enterobacter aerogenes ATCC 15038 (IFO 12010) and Pseudomonas sp. 4ASW, strains known to utilize a range of phosphonates as sole phosphorus source. In vitro phosphonatase activity was detected in extracts of both organisms; however extensive analysis failed to detect any organic product from phosphonates other than phosphonoacetal dehyde. Non-specific liberation of phosphate was observed in Pseudomonas sp. 4ASW, associated with a single fraction of FPLC-purified extract, and is believed to result from the activity of cellular phosphatases.

Chromatography, High Pressure Liquid↗
PubMed1991

Purification and characterization of an oxygen-insensitive NAD(P)H nitroreductase from Enterobacter cloacae.

The reductive products of several nitroaromatic compounds have been found to be toxic, mutagenic, and carcinogenic. The nitroreductases present in intestinal microflora have been implicated in the biotransformation of these compounds to their deleterious metabolites. A "classical" nitroreductase has been purified from Enterobacter cloacae 587-fold using a protocol which yields approximately 1 mg of purified nitroreductase from 10 liters of cell culture. An analysis of the physical properties of the nitroreductase indicates that the enzyme is active as a monomer with a calculated molecular mass of 27 kDa. FMN has been identified as a required flavin cofactor and is present at a stoichiometry of 0.88 mol of FMN bound/mol of active enzyme. The enzyme was found capable of reducing nitrofurazone under aerobic conditions indicating that the mechanism involves an obligatory two-electron transfer. Thus, this enzyme can be classified as an oxygen-insensitive nitroreductase. The purified nitroreductase can utilize either NADH or NADPH as a source of reducing equivalents and can reduce a variety of nitroaromatic compounds including nitrofurans and nitrobenzenes as well as quinones. Studies in which the rates of nitroreduction for a series of para substituted nitrobenzene derivatives were determined suggest that a linear free energy relationship exists between the rate and the redox midpoint potential of the substrate.

Bacterial Proteins↗
PubMed1991

Cloning, nucleotide sequence, and expression of the nitroreductase gene from Enterobacter cloacae.

The "classical" nitroreductases of enteric bacteria are flavoproteins which catalyze the reduction of a variety of nitroaromatic compounds to metabolites which are highly toxic, mutagenic, or carcinogenic. The gene for the nitroreductase Enterobacter cloacae has now been cloned using an antibody specific to this protein. The nucleotide sequence of the structural gene and flanking regions are reported. Sequence analysis indicates that this gene belongs to a gene family of flavoproteins which have not been previously described. Analysis of the 5'-untranslated region reveals the presence of putative regulatory elements which may be involved in the modulation of the expression of this enzyme. The cloned gene was placed under the control of a T7 promoter for overexpression of the protein in Escherichia coli. The expressed recombinant protein was purified to homogeneity and exhibited physical, spectral, and catalytic properties identical to the protein isolated from E. cloacae.

Amino Acid Sequence↗
PubMed1991

Pyridoxal phosphate-dependent histidine decarboxylases. Cloning, sequencing, and expression of genes from Klebsiella planticola and Enterobacter aerogenes and properties of the overexpressed enzymes.

The hdc genes encoding the inducible pyridoxal-P-dependent histidine decarboxylase (HisDCase) of Klebsiella planticola and Enterobacter aerogenes were isolated, sequenced, and expressed in Escherichia coli under control of the lac promoter, and the overproduced enzymes were purified to homogeneity from the recombinant host. Formation of inclusion bodies during synthesis of the E. aerogenes enzyme was avoided by cooling the culture and inducing at 25 degrees C. The cloned enzymes were produced in amounts three to four times those present in the fully induced native hosts and were identical in properties to those isolated earlier (Guirard, B. M., and Snell, E. E. (1987) J. Bacteriol. 169, 3963-3968). The two enzymes showed 85% sequence identity and also showed 80% sequence identity with the previously sequenced (Vaaler, G. L., Brasch, M. A., and Snell, E. E. (1986) J. Biol. Chem. 261, 11010-11014) HisDCase of Morganella morganii. Nevertheless, antibodies to the M. morganii HisDCase do not cross-react with these enzymes suggesting that the regions of amino acid variations are located on the outer surface of the proteins. All three HisDCases are the same length (377 amino acid residues); encoded N-terminal methionine was completely removed in each case. These closely related pyridoxal-P enzymes show no sequence homology with the pyruvoyl-dependent HisDCases of Gram-positive bacteria.

Amino Acid Sequence↗
PubMed1990

[Neonatal meningitis caused by Enterobacter sakazakii: milk powder is not sterile and bacteria like milk too!].

Powdered milk for infants can contain very low numbers of Enterobacter sakazakii. Larger amounts of this organism can result in non-infective colonization. In infants, particularly the premature newborn, such colonization has been associated with abdominal distention and bloody diarrhoea or bacteriuria, but cases of sepsis and meningitis have also been reported. Infection has been associated with the use of contaminated spoons or blenders as well as the habit of keeping the ready-made milk hot in bottle-heaters. The risk of contamination of the milk can be eliminated by boiling bottles, teats and spoons as well as disinfecting the blender before use. The possibility of bacterial replication can be significantly reduced by keeping the ready-made milk in a refrigerator and warming it up immediately before use eg by using a microwave oven.

Enterobacter↗
PubMed1989

[Human infections caused by Enterobacter sakazakii. Microbiologic considerations].

The clinical and microbiological features of four patients with positive cultures for Enterobacter sakazaii are reported. They corresponded to two wound exudates, one appendicitis and one newborn conjunctivitis. The biochemical and physiological characteristics of this organism are analyzed (inability to ferment d-sorbitol and production of a non-diffusible yellow pigment); its difficult identification and the clinical rarity of its isolation are discussed. The relevant literature is reviewed.

Adult↗
PubMed1989

[Enterobacter, Serratia, Acinetobacter or Pseudomonas septicemias].

During 1985 and 1986, 48 bacteremias due to Enterobacter spp, Acinetobacter spp, Serratia spp or Pseudomonas spp were collected by the SES Group. The mortality rate of those infections due to multiresistant organisms appeared high (27.9%). Among all factors influencing the prognosis, the following, age over 75 years, hospital-acquired infections, occurrence of shock and single antimicrobial therapy, are those associated with a high fatality rate. Given that clinical signs are not specific, nosocomial bacteremias should be treated by antimicrobial agents having wide spectrum, rapid onset of action and most often in combination.

Acinetobacter Infections↗
PubMed1989

Isolation and survival of gentamicin resistant Enterobacter aerogenes on finger tips of hospital personnel.

Enterobacter aerogenes was isolated from the finger tips of 13.3 percent of hospital personnel while they were working in the wards. Gentamicin resistant strains were isolated more frequently than gentamicin sensitive. Carriage rate of E. aerogenes was higher among nurses than other staff members. There was no correlation between antibiotic resistance, capsular serotypes and survival of these strains on finger tips. The property of prolonged survival on finger tips is chromosomal in nature and not mediated by conjugative or non-conjugative plasmids.

Cross Infection↗
PubMed1989

[Meconium peritonitis complicated with Enterobacter aerogenes infection--a case report].

Meconium peritonitis is an aseptic peritonitis that follows a perforation of the intestines in fetal life. It is a serious complication of intestinal perforation. This article reports a female newborn who was born with gestational age of 40 weeks and birth body weight of 3400 gm. She was admitted at age of 6 days, owing to persistent vomiting, abdominal distention, tachypnea and acrocyanosis since birth. The plain abdomen showed intra-abdominal calcification and intestinal distention. Under the impression of meconium peritonitis, she was treated aggressively with exploratory laparotomy, TPN and antibiotics. During operation, dense fibrous adhesions and agglutination of the intestine were found. But no intestinal perforation was noted. Culture of ascites isolated Enterobacter aerogenes. The patient's condition was gradually recovered and she was discharged at age of 32 days. According to the previous literature, meconium peritonitis is a relative rare disease with poor prognosis. So we would like to report this case.

Enterobacter↗
PubMed1989

[Nosocomial infections with resistant Enterobacter cloacae. Effects on the disease and the treatment].

We describe five patients with serious nosocomial infection caused by resistant Enterobacter cloacae. Four of the patients underwent cardiovascular operations and were infected with an identical, multiple beta-lactam resistant E cloacae strain. The fifth patient was admitted with several wounds and skin-ulcers containing both resistant and sensitive E cloacae of other types. The resistant strain caused septicemia in four patients; three developed multiorgan failure and one died. The most probable causes of these infections were an abundant and constant reservoir of E cloacae in a sink at the operating unit, periodical breakdown of barriere routines, and heavy use of cephalosporins.

Adult↗
PubMed1986

[Failure of new beta-lactam antibiotics in the treatment of severe Enterobacter cloacae infections].

In 12 patients infected with an Enterobacter cloacae (E. cl.) initially susceptible to the 3rd generation cephalosporins, we observed the emergence in vivo of variants resistant to most of the new beta-lactam antibiotics (carboxy-penicillins, ureido-penicillins, 3rd generation cephalosporins and aztreonam). These variants remained susceptible to mecillinam and imipenem. The variant emerged under treatment with cefotaxime in 3 cases, with moxalactam in 3 cases, with aztreonam, carbenicillin, and ticarcillin in 1 case each and without treatment in 1 case. An aminoglycoside was combined with the beta-lactam antibiotic in 6 cases. Therapeutic failure was attributed to emergence of the resistant variant in 6 out of 12 cases (with an aminoglycoside in 3 cases, with the beta-lactam antibiotic alone in 3 cases). These case reports underline the importance of bacteriological monitoring of patients infected with E. cl. treated with a beta-lactam antibiotic. The susceptibility to beta-lactam antibiotics of the E. cl. strains isolated during treatment should be systematically retested.

Adult↗
PubMed1988

[Action of imipenem on Enterobacter cloacae].

Most of Enterobacter cloacae strains produce chromosomally determined class I beta-lactamases when they are exposed to beta-lactams. Imipenem is a strong inducer of these enzymes but is poorly affected by them. We compared the effect of imipenem on inducible, non-inducible and stably derepressed strains of E. cloacae using the killing curve system. With antibiotic concentrations of 0.5 mg/l or more, an intense dose-dependent bactericidal effect was observed within 4 to 6 hours. However the bactericidal activity was incomplete. With an inoculum as low as 10(5) bacteria/ml a regrowth was observed after 24 hours. Among the surviving bacteria imipenem had the same bactericidal kinetic than on the parental strain. Similar results were observed with inducible, non-inducible and stably derepressed strains.

Cefotaxime↗
PubMed1987

Ciprofloxacin inhibition of cefoxitin betalactamase induction in an Enterobacter cloacae strain.

The betalactamase activity of an Enterobacter cloacae strain was measured using subinhibitory concentrations of cefoxitin and ciprofloxacin respectively as enzyme inducer and induction inhibitor. Cultures of this strain were also exposed to different ceftriaxone concentrations either alone or in the presence of cefoxitin and/or ciprofloxacin. Ciprofloxacin at subinhibitory concentrations does not interfere with ceftriaxone's activity and is capable of neutralizing the antagonistic effect of cefoxitin on ceftriaxone. The enzyme assay and MS-2 System demonstrate ciprofloxacin's partial suppression of cefoxitin betalactamase induction in the E. cloacae strain tested.

Cefoxitin↗
PubMed1987

Intracellular bioaccumulation of zinc by an Enterobacter species.

Bacterial strains tolerant to the presence of 100 mg/l zinc ions were isolated from a water reclamation system. Each of the organisms were screened for their ability to accumulate zinc at the above mentioned concentration. The organism capable of maximum zinc accumulation was identified as an Enterobacter species and intracellular zinc deposition by this micro-organism was determined using energy dispersive X-ray analysis, transmission electron microscopy and a metal-staining technique at the light microscopy level. Further studies revealed that growth and glucose utilization by this isolate were inhibited in the presence of zinc, compared to a control culture grown in the absence of zinc.

Biological Transport↗