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Biomedical subjects

C K Kim

Publications and source records attributed to C K Kim.

At least 145 records · Page 8Linked to original sources
PubMed1998

Changes in serum haptoglobin level after allergen challenge test in asthmatic children.

Bronchial asthma is characterized by airway inflammation, which underlies the phenomenon of bronchial hyperresponsiveness. Previous studies have shown that this correlates with the serum concentration of haptoglobin. The occurrence of the late asthmatic response (LAR) after an allergen challenge test is associated with airway inflammation. The objectives of this study were to examine serum levels of haptoglobin during the 24 h after allergen challenge and to compare changes between the subjects with and without LAR. We studied two groups of children with perennial asthma who developed the early asthmatic response (EAR) only (group I: n = 14), and EAR but also LAR (group II: n = 14) after an allergen (Dermatophagoides pteronyssinus) challenge test. Serum concentrations of haptoglobin were measured at baseline, at EAR, and at 2 h (recovery), 8 h (LAR), and 24 h after the challenge. Baseline levels were similar in the two groups (group I: 128 +/- 57 mg/dl; group II: 129 +/- 50 mg/dl). In group I, there was no significant change in the level at any time point; in contrast, the subjects in group II showed a relative fall (92 +/- 40 mg/dl) at 8 h, and an increase (161 +/- 79 mg/dl) at 24 h after the challenge. Our results indicate that the serum concentration of haptoglobin decreases at the time of LAR and is subsequently replenished during the ensuing time. Although further studies are needed, we think that haptoglobin may be inflused into the airways during the inflammatory process associated with LAR, and that this may be followed by "overshooting" production.

Allergens↗
PubMed1998

Serum levels of soluble interleukin-2 receptor at acute asthma exacerbation: relationship with severity of exacerbation and bronchodilator response.

BACKGROUND: T lymphocytes play a central role in the regulation of airway inflammation in asthma, and T cell activation appears to be a characteristic feature of acute asthma. It is not clear, however, whether this is proportional to the severity of acute asthma and is directly related to airway inflammation relevant to airflow obstruction during acute asthma. It is presumed that the extent to which bronchoconstriction or inflammation contributes to airflow obstruction in acute asthma may determine responsiveness to bronchodilator therapy. METHODS: Fifty asthmatic children who visited the emergency room due to acute exacerbation were studied. Serum levels of soluble interleukin-2 receptor (sIL-2R), a marker of T cell activation, were measured at the time of acute exacerbation and of clinical remission. At acute exacerbation, FEV1 was assessed before and after the administration of aerosolized salbutamol. RESULTS: The mean (+/-SD) serum level of sIL-2R at acute exacerbation (854+/-248 U/ml) was significantly higher (p<0.01) than at clinical remission (676+/-211 U/ml). It correlated positively with the severity of exacerbation (r = 0.47, p<0.01), but showed no significant relationship with bronchodilator response (r = -0.17, p = 0.20). CONCLUSION: A higher serum level of sIL-2R at acute exacerbation was associated with more severe exacerbation but not with lower bronchodilator response. These findings suggest that in the context of acute asthma, T cell activation is proportional to disease activity, but its relationship to airway inflammation relevant to the genesis of airflow obstruction remains obscure.

Acute Disease↗
PubMed1998

Periodontal repair in dogs: effect of allogenic freeze-dried demineralized bone matrix implants on alveolar bone and cementum regeneration.

The objective of this study was to evaluate alveolar bone and cementum regeneration following surgical placement of an allogenic, freeze-dried, demineralized bone matrix (DBM) cortical strip implant. Critical size, supraalveolar periodontal defects were surgically created around the second, third, and fourth mandibular premolar teeth in eight mongrel dogs. Contralateral jaw quadrants in six animals were randomly assigned to receive the DBM implant, or serve as surgical control. Two additional animals received bilateral DBM implants. Flaps were coronally advanced to submerge teeth and implants, and sutured. Three animals were exited from the study due to extensive early wound failure. Remaining animals were sacrificed at 8 weeks postsurgery. Histometric recordings included defect height, bone regeneration/DBM implant height, cementum regeneration height, root resorption, and ankylosis. Large areas of unresorbed DBM exhibiting fragmentation and empty osteocyte lacunae were observed adjacent to new bone formation, or bone formation was observed adjacent to or within the implant, often exhibiting ankylosis. Cementum regeneration appeared enhanced in shelter of the DBM implant. Histometric recordings (mean+/-SD) for DBM and control defects, respectively, were: defect height, 4.8+/-0.2 mm and 4.4+/-0.2 mm; bone regeneration/DBM implant height, 4.0+/-1.3 mm and 1.2+/-0.6 mm; cementum regeneration height, 1.4+/-0.4 mm and 0.7+/-0.2 mm; root resorption, 0.5+/-0.3 mm and 1.2+/-0.3 mm; and ankylosis, 0.5+/-0.2 mm and 0.1+/-0.1 mm without statistically significant differences between experimental conditions (N=3). Within the limitations of this study, the histologic observations suggest that surgical implantation of allogenic, freeze-dried DBM cortical strip implants may have a potential to support cementum regeneration, possibly by providing conditions for guided tissue regeneration, however, alveolar regeneration appears unpredictable.

Alveolar Bone Loss↗
PubMed1998

Periodontal repair in intrabony defects treated with a calcium sulfate implant and calcium sulfate barrier.

THIS RANDOMIZED, CONTROLLED, CLINICAL STUDY was designed to evaluate outcome following surgical implantation of an allogeneic, freeze-dried, demineralized bone matrix-calcium sulfate (DBM+CS) composite with a CS barrier in intrabony periodontal defects. Twenty-six patients contributing 26 deep intrabony defects completed the study. Thirteen patients received the DBM+CS implant. Thirteen patients received gingival flap surgery alone (GFS; control). Clinical outcome was assessed at 6 and 12 months postsurgery. At 12 months postsurgery, probing depth (PD) reduction (mean +/-SD) for the DBM+CS and GFS group was to 4.3+/-0.5 and 3.0+/-1.3 mm; clinical attachment gain was to 2.9+/-0.8 and 1.7+/-1.5 mm; and probing bone level gain was to 2.9+/-1.4 and 1.2+/-1.2 mm, respectively. There were no apparent differences between evaluations at 6 and 12 months postsurgery. Clinical improvements were significantly different from presurgery for both groups at both observation intervals (P < 0.01). There were no significant differences between groups in PD reduction and clinical attachment gain. Probing bone level gain was significantly greater in the DBM+CS group compared to controls (P < 0.05). In summary, surgical implantation of DBM+CS with a CS barrier resulted in reduced PD and improved attachment levels comparable to that achieved by gingival flap surgery alone. However, gain in probing bone levels in deep intrabony periodontal pockets assessed by clinical parameters was greater than that observed by gingival flap surgery alone. These changes were noted at both 6 and 12 months after surgery. This regenerative technique needs further biologic evaluation before being generally accepted.

Adult↗
PubMed1998

Effect of a calcium sulfate implant with calcium sulfate barrier on periodontal healing in 3-wall intrabony defects in dogs.

This controlled, split-mouth, preclinical study was designed to evaluate outcome following surgical implantation of an allogeneic, freeze-dried demineralized bone matrix-calcium sulfate (DBM+CS) composite with a CS barrier in 3-wall intrabony periodontal defects in 4 dogs. Control conditions included surgical implantation of DBM or CS and gingival flap surgery (GFS) alone. Three-wall intrabony defects (4x4x4 mm) were surgically created at the mesial and distal aspect of the maxillary and mandibular first and third premolars, respectively. Maxillary and mandibular defects each received 1 of the 4 experimental conditions. Experimental conditions were rotated between defect sites in subsequent animals. Block sections of the defects were collected at sacrifice 8 weeks postsurgery and processed for histometric analysis. Histometric defect height (means +/- SD) for the DBM+CS, DBM, CS, and GFS groups amounted to 4.2 +/- 0.5, 4.3 +/- 0.7, 4.0 +/- 0.2, and 4.1 +/- 0.2 mm, respectively. Connective tissue adhesion (connective tissue contact to the root without apparent cementum formation) amounted to 0.4 +/- 0.3, 0.4 +/- 0.3, 0.5 +/- 0.2, and 1.6 +/- 0.5 mm for the DBM+CS, DBM, CS, and GFS groups, respectively; the DBM+CS, DBM, and CS groups being significantly different from the GFS group (P < 0.05). Cementum regeneration amounted to 3.0 +/- 0.3, 3.1 +/- 0.4, 2.5 +/- 0.4, and 1.6 +/- 0.3 mm for the DBM+CS, DBM, CS, and GFS groups, respectively; the DBM+CS, DBM, and CS groups being significantly different from the GFS group (P < 0.05). Alveolar bone regeneration amounted to 2.7 +/- 0.4, 2.7 +/- 0.3, 1.8 +/- 0.5, and 0.7 +/- 0.1 mm for the DBM+CS, DBM, CS, and GFS groups, respectively; the DBM+CS, DBM, and CS groups being different from the GFS group (P < 0.05), and the DBM+CS and DBM groups being different from the CS group (P < 0.05). None of the DBM-containing implants provided evidence of bone metabolic activity. In summary, surgical implantation of DBM and CS, alone or in combination, may result in significantly improved regeneration of alveolar bone and cementum in this preclinical model. Observed regeneration is likely unrelated to a biologic activity inherent in DBM. Rather it appears that space-providing properties of the implants supported observed regeneration.

Alveolar Bone Loss↗
PubMed1998

Enhancement of bioavailability of ketoprofen using dry elixir as a novel dosage form.

To enhance the dissolution rate and bioavailability of poorly water-soluble ketoprofen, a novel oral dosage form of ketoprofen, termed ketoprofen dry elixir, was developed by the spray-drying technique. Ketoprofen, dextrin, and sodium lauryl sulfate were dissolved in an ethanol-water mixture (20:25 w/w) and thereafter spray-dried to form the ketoprofen dry elixir. Comparative studies on the in vitro dissolution and in vivo adsorption of ketoprofen in the form of dry elixir and powder were carried out. Ketoprofen in the dry elixir completely dissolved within 5 min. On the other hand, only about 50.1% of ketoprofen powder alone dissolved during 60 min. The initial dissolution rate of ketoprofen in the dry elixir markedly increased in distilled water at 37 degrees C, becoming fourfold higher than that of ketoprofen powder alone. The maximal plasma concentration of ketoprofen (Cmax) and the area under the concentration-time curve from zero to 8 hr (AUC0-8 hr) after the oral administration of dry elixir increased about 3.2- (24.6 versus 7.6 micrograms/ml) and 2.2-(38.4 versus 17.3 micrograms hr/ml) fold compared with powder alone. It was obvious that ketoprofen dry elixir might be a useful solid dosage form to improve the dissolution rate and bioavailability of poorly water-soluble ketoprofen.

Administration, Oral↗
PubMed1998

Prolonged release of tegafur from S/O/W multiple emulsion.

To develop a prolonged and sustained release preparation, we prepared an albumin microsphere-in-oil-in-water emulsion (S/O/W) and examined sustained release from it in comparison with other control preparations such as water-in-oil (W/O) emulsions and microspheres in vitro and in vivo, respectively. Tegafur was used as a model drug. A microsphere-in-oil emulsion was prepared by adding albumin microspheres to soybean oil containing 20% Span 80. To prepare an S/O/W emulsion, the microsphere-in-oil emulsion was added into an aqueous solution of hydroxypropyl methylcellulose containing Pluronic F68. The mean particle size of the albumin microspheres was 3 microns, and the ratio of entrapment of tegafur into albumin microspheres was about 25%. In an in vitro release test, the t75 of the S/O/W emulsion was fourfold greater and in an in vivo release test the mean residence time of tegafur from the S/O/W emulsion was more than twofold that from a W/O emulsion or microsphere system. The mean residence time of 5-fluorouracil (5-FU) from an S/O/W emulsion was also greater than with other dosage forms. These results suggest the possible usefulness of an S/O/W emulsion for the sustained and prolonged release of tegafur.

Animals↗
PubMed1998

A case of CMV disease of the jejunum in a patient with non-Hodgkin's lymphoma.

CMV infection may occur anywhere in the gastrointestinal tract. Among the small intestine, ileum is the most common site of CMV disease and infection of jejunum is a rare one in patients with CMV gastroenteritis. Although rare, the reason why the recognition of this diagnosis is important is that it cause the lethal hemorrhage and perforation of gastrointestinal tract when its diagnosis and treatment was delayed. Rapid diagnosis are able to using the immunohistochemical stain in shell vial culture of infected specimen or peripheral neutrophils preparation in viremic patients within 8 to 36 hours. The treatment of choice is antiviral agent or surgical resection. We experienced a case of CMV disease of jejunum in patient with non-Hodgkin's lymphoma who showed severe ulceration in jejunum and massive intestinal hemorrhage, and he survived after successful treatment with segmental resection of jejunum and intravenous ganciclovir.

Adult↗
PubMed1998

Prenatal ethanol exposure differentially alters behavior in males and females on the elevated plus maze.

Rodents prenatally exposed to ethanol demonstrate hypothalamic-pituitary-adrenal and behavioral hyperactivity to a variety of stressful situations. The present study examined both behavioral and corticosterone (CORT) responses to the elevated plus maze (+-maze), an anxiety- or fear-provoking task. Sprague-Dawley male and female offspring from fetal ethanol-exposed (E), pair-fed (PF), and ad libitum-fed control (C) groups were tested at 60 to 90 days of age. In experiment 1, behavior was measured in animals exposed to the +-maze for 5 min on two consecutive days; 2 weeks later, both behavioral and CORT responses were measured in animals confined to the open and closed arms of the maze for 20 min. In experiment 2, animals were placed in an open field (OF) for 5 min before a single 5-min exposure to the +-maze. Factor analysis of the scored behaviors from the two experiments indicated two main factors, designated "exploration" and "fear." E males and females both exhibited higher levels of exploratory behaviors when placed directly on the +-maze from their homecages without prior exposure to the OF, compared with C males and females. In addition, when confined to the closed arms of the +-maze, E males and females demonstrated higher levels of activity, compared with C males and females. After OF exposure, however, both E males and females demonstrated lower levels of exploratory behaviors than C males and females, and E females also had increased CORT levels, compared with PF and C females. Interestingly, E females, but not E males, showed an increase in fear-related behaviors on the +-maze, compared with controls, regardless of prior OF exposure. These data demonstrate that prenatal ethanol exposure may differentially affect both behavioral and hormonal responses of males and females in an aversive behavioral task and suggest that there may be a sex difference in the sensitivity of the mechanism(s) underlying these responses.

Animals↗
PubMed1998

Influence of nitric oxide synthase inhibitors on the ACTH and cytokine responses to peripheral immune signals.

Nitric oxide (NO) is an unstable gas that participates in the response of the hypothalamic-pituitary-adrenal (HPA) axis to a variety of immune signals, including turpentine-induced tissue damage and the systemic injection of the pro-inflammatory cytokine interleukin 1-beta (IL-1beta). Studies that have investigated the role of this gas in the intact rat have relied on blockade of NO formation with the NO synthase (NOS) inhibitor N(omega)nitro-L-arginine-methylester (L-NAME). They have suggested that endogenous NO blunts the ACTH response to intravenous (i.v.) IL-1beta in part by exerting an inhibitory influence on the release of hypothalamic peptides such as corticotropin-releasing factor (CRF) from nerve terminals in the median eminence. It must nevertheless be noted that, at present, evidence for this mode of action remains circumstantial. Significant controversy remains regarding the specificity of the compounds used to block NO formation, the characteristics of their effect in terms of dose and timing of administration, the possibility that their effect is restricted to IL-1beta or can be expanded to other pro-inflammatory cytokines, and the question of whether the possibility that they might also influence ACTH release by altering circulating levels of tumor necrosis factor-alpha (TNF-alpha) and IL-6. The purpose of the present study was to elucidate these points. In the first series of experiments, we determined the i.v. IL-1beta-induced ACTH response to various doses of systematically injected L-NAME (1-100 mg/kg). At 10-100, but not 1 mg/kg, L-NAME significantly (P<0.01) augmented the ACTH response to IL-1beta, with a maximum effect observed at 30 and 100 mg/kg. At the 30 mg/kg dose, L-NAME was equally effective in augmenting the ACTH response when administered between 5 and 240 min prior to the cytokine. The effect of L-NAME was fully mimicked by equivalent doses of other arginine derivatives such as N(omega)-monomethyl-L-arginine (L-NMMA) or N(omega)-nitro-L-arginine (L-NNA), indicating that controversy in the published literature concerning the influence of NO on CRF secretion does not appear to be due to the use of different arginine derivatives. The ability of other cytokines such as TNF-alpha and IL-6 to release ACTH and corticosterone was significantly (P<0.01) augmented by blockade of NO formation in a manner similar to that found with IL-1beta. To test the hypothesis that L-NAME might alter ACTH secretion at least in part by modifying the secretion of pro-inflammatory cytokines, we measured plasma concentrations of TNF-alpha and IL-6 following endotoxin injection in the presence or absence of L-NAME. Blockade of NO formation reduced TNF-alpha but increased IL-6 levels in rats administered the lipopolysaccharide (25 microg/kg i.v.). As L-NAME augments the ACTH response to TNF-alpha as well as IL-6, it is improbable that changes in TNF-alpha and IL-6 secretion during immune stimulation represents an important mechanism mediating the inhibitory influence of endogenous NO on the HPA axis activity. Collectively, these results indicate that the systemic injection of L-NAME very quickly augments the stimulatory effect of pro-inflammatory cytokines on ACTH secretion, and does so for at least 4 h. Other arginine derivatives known to block the activity of constitutive NO syntheses, such as L-NMMA and L-NNA, exert an effect that is virtually identical to that of L-NAME. The ability of L-NAME to increase the ACTH response to i.v. IL-1beta is also observed in rats injected with TNF-alpha and IL-6. Because of the opposite effects of L-NAME on the levels of these two cytokines, the influence of arginine derivatives on ACTH release is probably not due to changes in cytokines produced during immune stimulation such as endotoxemia.

Adrenocorticotropic Hormone↗
PubMed1998

Thallium-201 retention in focal intracranial lesions for differential diagnosis of primary lymphoma and nonmalignant lesions in AIDS patients.

UNLABELLED: The purpose of this study was to determine whether 201Tl retention in focal intracranial lesions can help distinguish central nervous system (CNS) lymphoma from toxoplasmosis and other nonmalignant CNS lesions in patients with acquired immunodeficiency syndrome. METHODS: Forty-nine patients who presented with focal lesions on CT and/or MRI had 201Tl brain SPECT studies (early and delayed image sets) performed shortly after admission. Early and delayed 201Tl uptake ratios were obtained for the positive studies, and the retention index of thallium was calculated (delayed/early target-to-background mean count ratio). RESULTS: Twenty-nine patients had foci of significantly increased 201Tl uptake on the early images in regions of corresponding CT/MRI lesions. Ten of these patients had biopsy-proven lymphomas. Another patient was found to have metastatic adenocarcinoma. Twelve additional patients had a response to radiation therapy or a clinical course consistent with lymphoma and six patients had a false-positive SPECT study. The early uptake ratio could not separate malignant from nonmalignant lesions. The 201Tl retention index in patients with lymphomas (1.18 +/- 0.16) was significantly higher than the retention index in adenocarcinoma (0.24) and in the six nonmalignant lesions (0.62 +/- 0.07). The lowest retention index in patients with lymphoma was 1.07, and the highest retention index in nonmalignant lesions was 0.70. Twenty patients showed no 201Tl uptake in the regions of CT/MRI lesions. Three of them had biopsies consistent with a benign etiology, and one patient was diagnosed with tuberculosis. Fifteen patients improved clinically on antitoxoplasmosis medications alone, and one patient had CNS lymphoma. The overall sensitivity of 201Tl brain SPECT was 96%. The specificity was 76% by counting all studies with abnormal 201Tl uptake, but it increased to 100% when the retention index was also considered. CONCLUSION: The retention index increases the specificity of 201Tl brain SPECT in human immunodeficiency virus patients. In the presence of abnormal early 201Tl uptake, it is essential to perform delayed imaging and calculate the retention index to distinguish nonmalignant lesions from lymphoma. The absence of 201Tl uptake on early images at the site of a CT/MRI abnormality excludes the diagnosis of lymphoma with a high degree of confidence and delayed imaging is unnecessary.

AIDS-Related Opportunistic Infections↗
PubMed1998

Intramedullary fat necrosis of multiple bones associated with pancreatitis.

We describe findings of intramedullary fat necrosis on five imaging studies in a patient with alcoholic pancreatitis. Radiography and CT of extremities showed multiple osteolytic lesions that were initially considered to be metastases. However, a 99mTc-methylene diphosphonate whole-body bone scan revealed abnormal areas of increased uptake in only the bones of extremities without involvement of the axial skeleton, a distribution quite unusual for metastatic disease. Furthermore, 99mTc-sestamibi scintigraphy was essentially normal. MRI revealed findings compatible with the diagnosis of fat necrosis/infarct. Findings from bone biopsy demonstrated necrotic bone marrow without malignant cells. It may not be necessary to perform all the imaging studies described in this report when clinical features suggesting metastatic fat necrosis are present. Appearance and distribution of abnormalities on the whole-body bone scan and MR images show that necrosis/infarct of the marrow may obviate bone biopsy, which is often needed to confirm the diagnosis of intramedullary fat necrosis and to exclude neoplastic processes.

Aged↗
PubMed1997

Characterization of the gene encoding catechol 2,3-dioxygenase from Achromobacter xylosoxidans KF701.

Catechol 2,3-dioxygenase (C23O) catalyzes a meta cleavage of the aromatic ring in catechol to form 2-hydroxymuconic semialdehyde. A C23O gene was cloned from chromosomal DNA of A. xylosoxidans KF701, a soil bacterium degrading biphenyl, and expressed in E. coli HB101. In substrate specificity to catechol and its analogs, the C23O exhibited the highest aromatic ring-fission activity to catechol, and its relative activity to other dihydroxylated aromatics was 4-chlorocatechol > 4-methylcatechol > 3-methylcatechol >> 2, 3-dihydroxybiphenyl. Aromatic ring-fission activity of the C23O to catechol was about 40-fold higher than that to 2,3-dihydroxybiphenyl. Nucleotide sequence analysis of the C23O gene from A. xylosoxidans KF701 revealed an open reading frame consisting of 924 base pairs, and identified a putative ribosome-binding sequence (AGGTGA) at about 10 nucleotides upstream from the initiation codon. The open reading frame can encode a polypeptide chain with molecular weight of 34 kDa containing 307 amino acid residues. The deduced amino acid sequence of the C23O exhibited the highest homology with that of C23O from Pseudomonas sp. IC with 96% identity, and the least homology with that of C23O from P. putida F1 with 22% identity among reported C23O sequences. Furthermore, comparison of the C23O sequence with other extradiol dioxygenases has led to identification of evolutionally conserved amino acid residues whose possible catalytic and structural roles are proposed.

Alcaligenes↗
PubMed1997

Localization and sequence analysis of the phnH gene encoding 2-hydroxypent-2,4-dienoate hydratase in Pseudomonas sp. strain DJ77.

The phnDEFG genes of Pseudomonas sp. DJ77, which are responsible for the degradation of polyaromatic hydrocarbons and chlorinated aromatics, were located previously on the 6.8 kb XhoI fragment of chromosomal DNA. Here, we sequenced a downstream region hitherto unknown and identified the phnH gene encoding a 2-hydroxypent-2,4-dienoate hydratase, which is required for the conversion of 2-hydroxypent-2,4-dienoate to 4-hydroxy-2-oxovalerate in the meta-cleavage pathway of catechols. The relative position of the hydratase gene in the phn operon is unique compared to the other meta-cleavage operons which have a dehydrogenative branch of the pathway. The PhnH hydratase contains 264 amino acids with a Mr of 28048. The deduced amino acid sequence of the PhnH enzyme is 60.9-31.6% identical to those of homologous enzymes encoded by the todG, bphE, cmtF, bphH, bphX1, xylJ, dmpE, cumE, MTCY03C7.20 and etbE genes.

Amino Acid Sequence↗
PubMed1997

Structure of catechol 2,3-dioxygenase gene encoded in TOM plasmid of Pseudomonas cepacia G4.

The catechol 2,3-dioxygenase is an extradiol-type dioxygenase which cleaves the C-C bond at the meta position of catechol to form 2-hydroxymuconic semialdehyde. A catechol 2,3-dioxygenase gene (tomB) in the TOM plasmid of P. cepacia G4 has been cloned and its nucleotide sequence was analyzed. The enzyme gene consisted of 945 base pairs with an ATG initiation codon and a TGA termination codon which can encode a polypeptide of molecular weight 35 kDa containing 314 amino acid residues, and a putative ribosome-binding sequence was identified at approximately 10 nucleotides upstream of the initiation codon. The deduced amino acid sequence of catechol 2,3-dioxygenase from P. cepacia G4 exhibited 79-82% homologies with those of 3-methylcatechol 2,3-dioxygenase of P. putida UCC2 and catechol 2,3-dioxygenase of P. pickettii PKO1.

Amino Acid Sequence↗
PubMed1997

Homogeneous liposome immunoassay for insulin using phospholipase C from Clostridium perfringens.

A new homogeneous liposome immunoassay system was developed in which analyte-phospholipase C conjugates are used instead of complement or melittin. This system was applied for the determination of insulin. Liposomes incorporated with calcein as a marker were prepared by the reverse-phase evaporation method. The lysis of liposomes was determined by measuring the fluorescence intensity of calcein released from liposomes and it was increased with increasing concentration of phospholipase C. Insulin was conjugated to phospholipase C by a three-step procedure with hetero-bifunctional crosslinking reagents, succinimidyl 4-(N-maleimidomethyl)cyclohexane-1-carboxylate and 3-(2-pyridyldithio)propionic acid N-hydroxysuccinimide ester. The lytic activity of phospholipase C was not affected by the reaction for conjugation. Both p-nitrophenylphosphatidylcholine hydrolytic activity and liposome lytic activity of insulin-phospholipase C conjugate were inhibited in the presence of insulin antiserum. However, lower concentration of conjugate and shorter incubation time were required when liposomes were used in the inhibition study. The antibody inhibition of conjugate-induced lysis could be reversed by addition of a competing amount of free insulin. The standard calibration curve was obtained in the range between 4 and 130 microIU/ml (r = 0.994). The detection limit (8 microIU/ml) was comparable with those of conventional heterogeneous enzyme immunoassays. This new assay approach offers a simple, sensitive, and inexpensive testing procedure for determining ultratrace amounts of bioactive substances.

Clostridium perfringens↗
PubMed1997

Tn5 tagging of the phenol-degrading gene on the chromosome of Pseudomonas putida.

Transposon mutagenesis was performed by the method of conjugational transfer in order to identify and characterize genes encoding enzymes involved in the pathway of phenol utilization as a carbon source. Escherichia coli, which carries the Tn5-132, Was mated with Pseudomonas putida SM25 as a host. We selected a mutant that could not utilize phenol as a carbon source. Chromosomal integration of the transposon was confirmed by Southern analysis, successfully tagging the gene related to a phenol-utilizing pathway. By cell-free enzyme and genetic complementation assays, the inactivated enzyme through the mutation of the corresponding gene was identified as the catB gene, which encodes a cis,cis-muconate lactonizing enzyme.

Biodegradation, Environmental↗