Biomedical subjects

K Kim

Publications and source records attributed to K Kim.

At least 271 records · Page 15Linked to original sources

PubMed1998

Catalytic center of DNA polymerase beta for excision of deoxyribose phosphate groups.

The amino-terminal 8-kDa domain of vertebrate DNA polymerase beta (pol beta) has an activity to excise deoxyribose phosphate (dRP) groups from 5'-incised apurinic/apyrimidinic (AP) sites during base excision repair. The excision reaction proceeds via a beta-elimination reaction following formation of a Schiff base between an aldehyde group of the AP site and an amino group of the enzyme. Here we report that the Lys-72 residue of this enzyme is the catalytic center for dRP excision. Substitutions of Lys-72 with Arg or Gln reduced the dRP excision activity to less than 1% of the wild-type 8-kDa domain, while substitutions of Lys-35, Lys-68, or Lys-84 did not abolish its activity. The Lys-72 mutations also significantly decreased Schiff base intermediates trapped by reduction with sodium borohydride. The 8-kDa domain alone was able to bind preferentially to a single-nucleotide gap or 5'-incised synthetic AP site on double-stranded DNA. The Lys-72 mutations did not affect this damage-specific DNA binding activity. When introduced into the intact enzyme, a mutation of Lys-72 to Arg did not affect DNA synthesis activity of pol beta, but eliminated the repair activity. Addition of the wild-type 8-kDa domain to this reaction restored the repair activity. These results indicate a specific role of Lys-72 of pol beta in the dRP excision during base excision repair.

Amino Acid Sequence↗

PubMed1998

The C2 domains of Rabphilin3A specifically bind phosphatidylinositol 4,5-bisphosphate containing vesicles in a Ca2+-dependent manner. In vitro characteristics and possible significance.

In the present study we investigated the lipid binding characteristics of the C2 domains of Rabphilin3a. We found that the tandem C2 domain of Rabphilin3a specifically bound lipid vesicles containing phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P2) in a Ca2+-dependent manner. There was little binding to vesicles containing PtdIns(3,4)P2 in the presence or absence of Ca2+. Binding to phosphatidylinositol 3,4,5-triphosphate-containing vesicles was similar to binding to PtdIns(4,5)P2-containing vesicles. The presence of physiological amounts of phosphatidylserine (PS) greatly potentiated the ability of PtdIns(4,5)P2 to cause vesicle binding. As with the C2 domains together, the binding of individual C2 domain of Rabphilin3a was much greater to PtdIns(4,5)P2-containing vesicles than PtdIns(3,4)P2-containing vesicles. Both C2 domains also bound 29 mol % PS-containing vesicles in a Ca2+-dependent manner. Because of the importance of the C2B domain in the enhancement of secretion from chromaffin cells by Rabphilin3a, its biochemistry was further investigated. The mutation of aspartates 657 and 659 to asparagines in C2B decreased Ca2+-dependent and increased Ca2+-independent vesicle binding, indicating the Ca2+ dependence of the domain is provided by aspartic acid residues in the putative Ca2+-binding pocket. A peptide from the COOH-terminal region of the C2B domain specifically inhibited ATP-dependent secretion from permeabilized chromaffin cells and the binding of Rabphilin3a to phosphatidylcholine/PS/PtdIns(4,5)P2-containing lipid vesicles, suggesting a role of this sequence in secretion through its ability to interact with acidic lipid vesicles.

Adaptor Proteins, Signal Transducing↗

PubMed1998

Involvement of flap endonuclease 1 in base excision DNA repair.

Base excision repair can proceed in either one of two alternative pathways: a DNA polymerase beta-dependent pathway and a proliferating cell nuclear antigen (PCNA)-dependent pathway. Excision of an apurinic/apyrimidinic (AP) site by cutting the phosphate backbone on its 3' side following incision at its 5' side by AP endonuclease is a prerequisite to completion of these repair pathways. Using a reconstituted system with the proteins derived from Xenopus laevis, we found that flap endonuclease 1 (FEN1) was a factor responsible for the excision of a 5'-incised AP site in the PCNA-dependent pathway. In this pathway, DNA synthesis was not required for the action of FEN1 in the presence of PCNA and a replication factor C-containing fraction. The polymerase beta-dependent pathway could also use FEN1 for excision of the synthetic AP sites, which were not susceptible to beta-elimination. In this pathway, FEN1 was functional without PCNA and replication factor C but required the DNA synthesis, which led to a flap structure formation.

Amino Acid Sequence↗

PubMed1998

Suppression of inflammatory responses by surfactin, a selective inhibitor of platelet cytosolic phospholipase A2.

Surfactin inhibits platelet and spleen cytosolic 100 kDa phospholipase A2 (PLA2). In contrast, this same compound enhances rat platelet group II PLA2 activity by approximately 2-fold and slightly increases group I PLA2 activity from porcine pancreas and Naja naja venom in vitro. Surfactin does not affect a Ca2+ -independent PLA2 partially purified from bovine brain. Thus, this compound inhibits selectively the cytosolic form of PLA2. Based on in vitro studies utilizing preincubation of surfactin with the enzyme, dialysis, and increased concentrations of substrates, the inhibitory effect of surfactin appears to be due to a direct interaction with the enzyme. Linear regression analysis of the linear portion of a concentration-response curve reveals an IC50 of 8.5 microM. To further determine the inhibitory pattern, a Dixon plot was constructed to show that the inhibition by surfactin is competitive, but not uncompetitive, with an inhibition constant of Ki = 4.7 microM in 50 mM Tris-HCl buffer, pH 8.0, at 37 degrees. Surfactin blocked non-stimulated and calcium ionophore A23187-stimulated release of arachidonic acid from monkey kidney CV-1 cells, which contain a cytosolic 100 kDa PLA2 as the major activity, as shown in an anionic exchange DEAE-5PW high performance liquid chromatography profile and western blotting analysis. Surfactin ameliorated inflammation induced by several chemicals. That is, it exhibited in vivo anti-inflammatory activity in several tested inflammatory reactions including 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced mouse ear edema, carrageenan-induced rat paw edema, and acetic acid-induced mouse writhing. These results demonstrate that surfactin is a selective inhibitor for cytosolic PLA2 and a putative anti-inflammatory agent through the inhibitory effect produced by direct interaction with cytosolic PLA2, and that inhibition of cytosolic PLA2 activity may suppress inflammatory responses.

Acetic Acid↗

PubMed1998

Mammalian peroxiredoxin isoforms can reduce hydrogen peroxide generated in response to growth factors and tumor necrosis factor-alpha.

Mammalian tissues express three immunologically distinct peroxiredoxin (Prx) proteins (Prx I, II, and III), which are the products of distinct genes. With the use of recombinant proteins Prx I, II, and III, all have now been shown to possess peroxidase activity and to rely on Trx as a source of reducing equivalents for the reduction of H2O2. Prx I and II are cytosolic proteins, whereas Prx III is localized in mitochondria. Transient overexpression of Prx I or II in cultured cells showed that they were able to eliminate the intracellular H2O2 generated in response to growth factors. Moreover, the activation of nuclear factor kappaB (NFkappaB) induced by extracellularly added H2O2 or tumor necrosis factor-alpha was blocked by overproduction of Prx II. These results suggest that, together with glutathione peroxidase and catalase, Prx enzymes likely play an important role in eliminating peroxides generated during metabolism. In addition, Prx I and II might participate in the signaling cascades of growth factors and tumor necrosis factor-alpha by regulating the intracellular concentration of H2O2.

Animals↗

PubMed1998

Ca2+-dependent protein kinases of Paramecium--cloning provides evidence of a multigene family.

Two genes for Ca2+-dependent protein kinases, PCaPK-alpha and PCaPK-beta, were isolated from a Paramecium genomic DNA library. The coding region of PCaPK-alpha encoded 481 amino acids and that of PCaPK-beta encoded 493 amino acids, predicting molecular masses of 55603 Da and 57131 Da for each putative protein. The sequences of the protein kinase catalytic domains of PCaPK-alpha and PCaPK-beta were closely related to those of the Ca2+-dependent protein kinases (CDPKs) from Plasmodium, Eimeria, and several plants, and the catalytic region of the Ca2+/calmodulin-dependent protein kinase family (35-48% identity). In the junction region between the catalytic and regulatory regions, only 9 of 31 amino acid residues are the same in the two Paramecium genes, and the sequences encoded in the Paramecium genes differ from those in the plant CDPK genes in about 20 of 31 residues in the junction region. The C-terminal region of the Paramecium kinases shared sequence similarity with Paramecium calmodulin (30-34% identity). Two Ca2+-dependent protein kinases previously characterized from Paramecium (52 kDa CaPK-1, and 50 kDa CaPK-2) are activated by Ca2+ in the micromolar concentration range and they directly bind Ca2+ in a 45Ca2+ overlay blot assay. The size predicted from the genes, the presence of four putative Ca2+-binding motifs encoded in PCaPK-alpha and PCaPK-beta, and the immunological cross-reaction of expressed cloned fragments of these genes with CaPK-2, suggest that they encode proteins of the same family.

Amino Acid Sequence↗

PubMed1998

Mutational analysis of domain II beta of bacteriophage Mu transposase: domains II alpha and II beta belong to different catalytic complementation groups.

This study examines the contribution of domain II beta of bacteriophage Mu transposase (A protein), a subdomain of the central catalytic domain II, to the transposition reaction. The properties of several point mutations implicate a role for this domain in facilitating metal-assisted assembly of the synaptic complex, as well as in intramolecular DNA strand transfer. Point mutations as well as deletions in domain II beta can be complemented by those in domain II alpha but not those in domain III alpha. Thus, residues within subdomains II alpha and II beta belong to different catalytic complementation groups.

Bacteriophage mu↗

PubMed1998

A randomized phase II trial of echinomycin, trimetrexate, and cisplatin plus etoposide in patients with metastatic nonsmall cell lung carcinoma: an Eastern Cooperative Oncology Group Study (E1587).

BACKGROUND: Patients with metastatic nonsmall cell lung carcinoma (NSCLC) usually have a poor prognosis. A chemotherapy regimen containing cisplatin is commonly used for symptom palliation. Echinomycin is a potent bifunctional intercalator of double-strand DNA; trimetrexate is a new derivative of methotrexate and is active against methotrexate-resistant tumor cells in vitro. METHODS: The Eastern Cooperative Oncology Group conducted a randomized Phase II study. Eligible patients were assigned to receive echinomycin 1200 microg/m2 by intravenous (i.v.) infusion over 30-60 minutes once a week for 4 weeks, repeated every 6 weeks; trimetrexate 12 mg/m2 i.v. bolus on Days 1-5 every 3 weeks, or 8 mg/m2 i.v. bolus on Days 1-5 for patients who had prior radiation to greater than 30% of their bone marrow; or cisplatin 60 mg/m2 i.v. on Day 1 and etoposide 120 mg/m2 i.v. on Days 1-3 every 4 weeks. Patients were evaluated before each cycle for tumor response, toxicity, and quality-of-life measurements. RESULTS: One hundred thirty-six patients were entered on the study, and 118 were evaluable for toxicity and response. The response rates were 16%, 5%, and 5% in patients treated with cisplatin and etoposide, echinomycin, and trimetrexate, respectively. There were no complete responses. The median survival was 37.9, 24.3, and 28.0 weeks for patients who received cisplatin and etoposide, echinomycin, and trimetrexate, respectively. Although cisplatin and etoposide appeared to give better therapeutic results, the response rate or survival did not reach statistical significance. This may have been due to inadequate sample size. Neither did quality-of-life measurement show any significant differences among treatments. CONCLUSIONS: Echinomycin and trimetrexate had minimal antitumor activity in patients with metastatic NSCLC: Response rate and survival remained poor in all three treatment arms. Patients should be encouraged to participate in clinical trials so that more effective therapy can be identified.

Adult↗

PubMed1998

The effect of organic solvents on the determination of cyclic boronates of some beta-blockers by gas chromatography/mass spectrometry.

Formation cyclic boronates for beta-blockers, by use of triethylamine (TEA) and pyridine as catalysts, gives more effective product yield. Eleven beta-blockers, formed by cyclic boronation with n-butylboronic acid and TEA, produced higher yields than by on-column thermal reaction and seven beta-blockers were shown to give the highest yields in the phenyl cyclic boronation with pyridine or TEA by on-column thermal and general reaction. The phenyl cyclic boronate of nadolol produced one peak in the chromatogram and the n-butyl cyclic boronate showed two peaks. On-column derivatization and n-butylboronic acid and pyridine was effective in saving analysis time and for convenience, even though some n-butyl cyclic boronates by cyclic boronation with TEA gave a better yield. In n-butyl cyclic boronation with pyridine by on-column thermal reaction the detection limits were 0.1 to 4 ng/microL in urine with a signal-to-noise ratio of 10:1.

Adrenergic beta-Antagonists↗

PubMed1998

Turn scanning by site-directed mutagenesis: application to the protein folding problem using the intestinal fatty acid binding protein.

We have systematically mutated residues located in turns between beta-strands of the intestinal fatty acid binding protein (IFABP), and a glycine in a half turn, to valine and have examined the stability, refolding rate constants and ligand dissociation constants for each mutant protein. IFABP is an almost all beta-sheet protein exhibiting a topology comprised of two five-stranded sheets surrounding a large cavity into which the fatty acid ligand binds. A glycine residue is located in seven of the eight turns between the antiparallel beta-strands and another in a half turn of a strand connecting the front and back sheets. Mutations in any of the three turns connecting the last four C-terminal strands slow the folding and decrease stability with the mutation between the last two strands slowing folding dramatically. These data suggest that interactions between the last four C-terminal strands are highly cooperative, perhaps triggered by an initial hydrophobic collapse. We suggest that this trigger is collapse of the highly hydrophobic cluster of amino acids in the D and E strands, a region previously shown to also affect the last stage of the folding process (Kim et al., 1997). Changing the glycine in the strand between the front and back sheets also results in a unstable, slow folding protein perhaps disrupting the D-E strand interactions. For most of the other turn mutations there was no apparent correlation between stability and refolding rate constants. In some turns, the interaction between strands, rather than the turn type, appears to be critical for folding while in others, turn formation itself appears to be a rate limiting step. Although there is no simple correlation between turn formation and folding kinetics, we propose that turn scanning by mutagenesis will be a useful tool for issues related to protein folding.

Carrier Proteins↗

PubMed1998

Childhood stress, behavioural symptoms and mother-daughter pubertal development.

Belsky, Steinberg and Draper (Child Development 1991, 62, 647-670) predicted that early childhood stress or conflict in the family environment would be associated with childhood behavioural symptoms, early puberty and early, less discriminate sexual behaviour. Their theory was tested in a retrospective self-report survey in 28 daughters (aged 12 to 15) and 21 mothers. In daughters, earlier menarche correlated with more family stress in a late childhood (age 7 to 11); more conflict with mother in early childhood (birth to age 6); more rejection from and less closeness to mother throughout childhood (birth to age 11); more anxiousness and internalizing symptoms (anxiousness/depression) in late childhood (age 7 to 11); earlier age at dating boys; and more boyfriends. An alternative interpretation is considered based on genetic transmission of maternal characteristics.

Adolescent↗

PubMed1998

Video-assisted thoracic surgery as a primary therapy for primary spontaneous pneumothorax. Decision making by the guideline of high-resolution computed tomography.

BACKGROUND: Because blebs are confirmed in most of the patients undergoing thoracotomy, identification of blebs by high-resolution computed tomography (HRCT) can be proposed as a surgical indication in primary spontaneous pneumothorax (PSP). If an apical bleb is identified, we treat the patient by video-assisted thoracic surgery (VATS). METHODS: From May 1995 to September 1997, 61 patients (21.9 +/- 4.6 years) were seen for initial episodes of PSP. Only seven showed bullae on simple chest radiography. However, by HRCT, 48 had sizable blebs (>5 mm), and 45 were treated surgically by VATS. RESULTS: The mean duration of chest tube use after surgery was 3.2 +/- 1.9 days, and the mean hospital stay was 4.5 +/- 1.9 days. Only one recurrence developed 5 weeks after VATS. CONCLUSIONS: Our protocol is effective in controlling an initial episode of PSP. It shortens the observation time before definitive surgical treatment, shortens the hospital stay, and decreases the likelihood of recurrence.

Adolescent↗

PubMed1998

Biliary cystadenoma of the liver.

Hepatobiliary cystadenoma is an uncommon lesion that is difficult to diagnose preoperatively. Here we report a 34-year-old woman who presented with enlargement of a cyst that had been observed for the previous 6 months. Diagnostic imaging revealed a 7-cm diameter cystic mass with irregular multiple septation in her liver. All laboratory test results were normal except for serum carbohydrate antigen (CA) 19-9 (62.5 U/ml). Because of the malignant potential and the history of enlargement, a complete surgical excision was performed. The patient was discharged after a good recovery; 2 months after surgery her serum CA19-9 level had returned to normal (32.9 U/ml). Regardless of the diagnostic modalities used, cystadenoma and cystadenocarcinoma cannot be differentiated with accuracy. Therefore complete surgical resection is the recommended therapy.

Adenoma, Bile Duct↗

PubMed1998

Surgical resection of mucoepidermoid carcinoma at the carina in a 9-year-old boy.

Mucoepidermoid carcinoma of trachea and bronchi is a rare tumor, especially in children. The authors report a case of 9-year-old boy with mucoepidermoid carcinoma at the carina. His presenting symptoms were hemoptysis of recent onset and intermittent cough of 2 years' duration. Preoperative assessment of the tumor was an intraluminal polypoid mass arising from the carina extending into the trachea and right main stem bronchus. A complete resection with reconstruction of carina was successful. The tumor was 12 mm in size, polypoid with a broad base. It had characteristic features of a low-grade mucoepidermoid tumor, namely, admixture of islands of intermediate cells and glandular components with invasion of submucosa. The patient is now 15 months postsurgery free of disease.

Bronchial Neoplasms↗

PubMed1998

Differential regulation of gonadotropin-releasing hormone (GnRH) receptor expression in the posterior mediobasal hypothalamus by steroid hormones: implication of GnRH neuronal activity.

The present study is designed to evaluate the relationship between gonadotropin-releasing hormone (GnRH) and GnRH receptor (GnRHR) gene expression during the steroid-induced LH surge. One week after ovariectomy (OVX), a capsule containing 17beta-estradiol (E) or vehicle (V) was implanted into OVX rats, and 2 days later a single injection of progesterone (P) or V was administered s.c. at 10:00 h. Poly(A)-rich RNA samples were isolated from the micropunches of the preoptic area (POA) and the posterior mediobasal hypothalamus (pMBH) from both sides of individual brain slices. Using competitive reverse transcription-polymerase chain reaction (RT-PCR) procedures, three parameters (POA GnRH, pMBH GnRHR) and pituitary GnRHR mRNA levels were simultaneously determined in each individual animal. POA GnRH mRNA and pituitary GnRHR mRNA levels were decreased by treatment with E, but increased by a combination of E and P. In contrast, pMBH GnRHR mRNA levels were clearly augmented by treatment with E, and decreased by the combination of E and P. Temporal changes in such parameters were determined in OVX+E+V- and OVX+E+P-treated rats. P augmented POA GnRH mRNA levels at the time of the LH surge (17:00 h) and the increased GnRH mRNA levels were remained until 22:00 h, while E alone failed to alter POA GnRH mRNA levels. In the pMBH micropunch samples, P substantially decreased E-induced increase in GnRHR mRNA levels at 17:00 h and further lowered those until 22:00 h. Antisense oligonucleotides of GnRHR mRNA administered into the lateral ventricle of OVX+E-treated rats blocked the E-induced increase in pMBH GnRHR mRNA levels. The antisense oligonucleotides also prevented the LH surge as well as the increase in pituitary GnRHR mRNA levels in the OVX+E+P-treated group. However, administration of this antisense oligonucleotides failed to alter POA GnRH mRNA levels. In conclusion, the present study demonstrated that there is an inverse relationship between POA GnRH mRNA levels and pMBH GnRHR mRNA levels in response to E and/or P, and that the blockade of the E-induced increase in pMBH GnRHR mRNA levels effectively nullified the P-induced LH surge. These results indicate that pMBH GnRHR gene expression is involved in synchronizing the GnRH neuronal activity, which is crucial for the generation of the LH surge.

Animals↗

PubMed1998

Retinoic acid regulates gonadotropin-releasing hormone (GnRH) release and gene expression in the rat hypothalamic fragments and GT1-1 neuronal cells in vitro.

The present study attempts to examine the possible involvement of retinoic acid (RA) in the regulation of gonadotropin-releasing hormone (GnRH) release and gene expression in the rat hypothalamic fragments and GT1-1 neuronal cells in vitro. During a short-term period (2h), RA (0.01-1 microM) increased GnRH release in a dose-related manner. Time-course experiments showed that RA rapidly increased GnRH release by 30 min in both cells. RA-induced GnRH release was slowly attenuated in the next incubation period in hypothalamic fragments, but rapidly returned to control levels in GT-1 cells. In hypothalamic fragments, GnRH mRNA levels decreased, but in GT1-1 cells, no change in GnRH mRNA levels was observed. We then extended the incubation time to see any changes in GnRH mRNA levels by RA in GT1-1 cells. In a long term (up to 48 h), RA increased GnRH mRNA levels in a dose- and time-related manner. Significant increase in GnRH mRNA levels by RA (at higher than 10 nM) was observed within 12h. Transient transfection experiments with a luciferase reporter vector containing more than 3 kb of the rat GnRH 5'-flanking region (-3002 to +88) revealed that RA also increased the rat GnRH promoter activity in a similar dose-and time-dependent manner, suggesting that increases in GnRH mRNA levels are attributable, at least in part, to the enhanced gene transcription. The promoter analysis with the 5'-deletional constructs demonstrated that cis-elements responsible for the RA action may reside within -1640/-1438 of the rat GnRH promoter, where multiple direct or palindromic arrangements of the AGGTCA-related sequences exist. We also showed that GT1-1 cells as well as the hypothalamic tissues express mRNA for multiple subtypes of retinoid receptors, and that reporter plasmids with three copies of the strong retinoic acid response element (RARE) were activated by 80 folds upon treatment with RA in GT1-1 cells, suggesting that retinoid receptors in GT1-1 cells are functional. Taken together, the present study strongly suggests that RA is an important regulator of the GnRH neurons.

Animals↗

PubMed1998

Open surgery for removal of a failing Gianturco stent with reversed sleeve resection of the right middle and lower lobes.

Although the use of a metallic stent in the treatment of benign tracheobronchial stenosis has been reported as a useful and safe technique, the incorporation of wire stents into the airway may be irreversible and is associated with problems. The authors' experience in a patient with incorrectly positioned metallic stent in the right main bronchus, which was successfully treated with bronchial sleeve resection, is presented.

Adult↗

PubMed1998

Response distribution as an explanation of the mirror effect.

Response distribution has recently been proposed as an explanation of the mirror effect in recognition memory. According to the proposal, participants presented with distinctive sets of items (e.g., low- and high-frequency words) vary their responses to give an equal number of positive responses (e.g., the sum of hits and false alarms) to each set. Four experiments tested this proposal. Two experiments showed that the mirror effect is present in the absence of distinctive sets of items. Two experiments showed that the mirror effects is present in the absence of response equalization. Wherever the response distribution hypothesis can be tested, it fails.

Adult↗