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Biomedical subjects

S Kim

Publications and source records attributed to S Kim.

At least 973 records · Page 54Linked to original sources
PubMed1994

Effect of enzymic methylation of heterogeneous ribonucleoprotein particle A1 on its nucleic-acid binding and controlled proteolysis.

Recombinant unmethylated heterogeneous nuclear ribonucleoprotein particle (hnRNP) protein A1 was enzymatically methylated by nuclear protein/histone protein methylase I [Rajpurohit, Lee, Park, Paik and Kim (1994) J. Biol. Chem. 269, 1057-1082] and the effect of methylation on several physiocochemical properties was studied. The relative binding-affinity of methylated and unmethylated protein A1 to nucleic acid was quite different. This was observed by the elution behaviour of the protein A1 on a single-stranded DNA/cellulose column; the concentration of NaCl required to release the bound protein A1 was 0.59 M for the methylated and 0.63 M for the unmethylated, respectively. Employing isoelectrofocusing, pI values of the methylated and unmethylated proteins were found to be 9.41 and 9.48, respectively. Maximum fluorescence quenching of protein A1 in the presence of coliphage MS2-RNA was found to be 40% with methylated and 45% with unmethylated. When both species of protein A1 were subjected to controlled trypsin digestion, t1/2 of the methylated protein was 1.31 min and the unmethylated, 1.63 min. The difference in their t1/2 values was much greater in the presence of MS2-RNA; 2.4 min for the former and 4.3 min for the latter, indicating that the methylated species was less stabilized by the RNA than the unmethylated. All of the above results consistently suggested that the binding-property of hnRNP protein A1 to single-stranded nucleic acid was significantly reduced subsequent to its arginine-methylation. The biological significance of this observation is discussed.

Arginine↗
PubMed1994

Mutational analysis of the substrate binding site of human complement factor D.

Complement factor D is a serine protease with a single natural substrate, C3b-complexed factor B, and very low catalytic activity against synthetic esters. The recently solved X-ray crystal structure of factor D has demonstrated certain key differences from other serine protease in the conformation of residues of the catalytic triad and the substrate-binding regions. To investigate possible contributions of unique amino acid substitutions to these distinct structural and functional features of factor D, we constructed a series of mutants by substituting trypsin substrate-binding residues for the corresponding factor D residues. Wild-type and seven mutant factor D cDNAs were expressed stably in Chinese hamster ovary cells, and the recombinant proteins were purified from culture supernatants and assayed by hemolytic, proteolytic, and esterolytic assays. The combined results indicate that residues Thr-198, Ser-199, Arg-202, and perhaps also Val-203 provide determinants for substrate binding and catalysis. The data also provide additional support for the hypothesis that the proteolytically active conformation of the active center of factor D is induced by its substrate, C3bB.

Amino Acid Sequence↗
PubMed1994

Renal vasodilating and diuretic actions of a selective endothelin ETB receptor agonist, IRL1620.

The effects of a selective agonist for endothelin ETB receptors, Suc-[Glu9,Ala11,15]endothelin-1-(8-21), IRL1620, on renal hemodynamics and urine formation were studied in anesthetized dogs. Intrarenal arterial infusion of IRL1620 at a dose of 50 ng/kg/min increased renal blood flow from 3.37 +/- 0.30 (mean +/- S.E.) to a maximal value of 4.43 +/- 0.45 ml/g kidney weight per min (ml/g/min) at 9.1 +/- 1.0 min after the start of infusion, with no change in systemic blood pressure and heart rate. Urine flow rate increased and urine osmolality, osmolar clearance and free water reabsorption decreased significantly whereas glomerular filtration rate remained unchanged. In dogs given ibuprofen (12.5 mg/kg, i.v.) after the start of infusion of the peptide, renal blood flow increased slightly but significantly from 3.78 +/- 0.82 to 4.17 +/- 0.96 ml/g/min (1.0 +/- 0.1 min), followed by a gradual reduction in renal blood flow. In dogs given L-NG-nitroarginine (75 micrograms/kg/min), the renal blood flow decreased following intrarenal administration of IRL1620 (50 ng/kg/min). It is suggested that IRL1620 enhances the release of nitric oxide and prostaglandins in the kidney and promotes renal vasodilation. The IRL1620-induced reduction of urine osmolality and free water reabsorption was affected by neither ibuprofen nor L-NG-nitroarginine, thereby suggesting that the suppression of urine concentration did not seem to be linked to the enhanced production of nitric oxide or prostaglandins in the kidney.

Animals↗
PubMed1994

Effect of adrenomedullin on renal hemodynamics and functions in dogs.

In order to elucidate the role of adrenomedullin in the kidney, we investigated the effects of adrenomedullin on renal hemodynamics and urine formation in anesthetized dogs. Intrarenal arterial infusion of adrenomedullin (0.8, 4 and 20 ng.kg-1.min-1) elicited dose-dependent increases in renal blood flow (by 10, 26 and 37%, respectively) with no change in blood pressure or heart rate, indicating a renal vasodilatory action of adrenomedullin. The glomerular filtration rate did not increase with the lower two doses, but increased marginally by 9% at the highest dose. Infusion of adrenomedullin at the rates of 4 and 20 ng.kg-1.min-1 increased urine flow and the urinary excretion of sodium and potassium dose dependently. Arterial and renal venous plasma renin activity was unaffected by adrenomedullin. These findings indicate that adrenomedullin is a potent renal vasodilatory peptide with a diuretic action. Since the threshold for the renal vasodilatory action of adrenomedullin is close to its physiological concentration in human plasma, adrenomedullin may play an important role in the regulation of renal function.

Adrenomedullin↗
PubMed1994

Role of angiotensin II in extracellular matrix and transforming growth factor-beta 1 expression in hypertensive rats.

The in vivo effects of alacepril (1-[(S)-3-acetylthio-2-methylpropanoyl]- L-prolyl-L-phenylalanine), an angiotensin converting enzyme inhibitor, and SC-52458 (5-[(3,5-dibutyl-1H-1,2,4-triazol-1- yl)methyl]-2-[2-(1H-tetrazol-5-ylphenyl)]pyridine), an angiotensin AT1 receptor antagonist, were examined on the cardiac and aortic gene expressions of extracellular matrices and TGF-beta 1 in young spontaneously hypertensive rats (SHR). In SHR, types I and III collagen mRNAs were increased in the left ventricle, and in contrast, fibronectin, collagen IV, and transforming growth factor-beta 1 (TGF-beta 1) mRNAs were increased in aorta, compared with those in Wistar-Kyoto rats. All the enhanced mRNAs in both organs in SHR were significantly inhibited by the short-term treatment with the above two drugs. Thus, angiotensin AT1 receptor may play an important role in the regulation of extracellular matrices and TGF-beta 1 expressions in SHR.

Angiotensin II↗
PubMed1994

An RNA binding site in a tRNA synthetase with a reduced set of amino acids.

A 30 amino acid helix-loop of known structure on the surface of the C-terminal domain of the class I Escherichia coli methionine tRNA synthetase is essential for methionine tRNA anticodon discrimination. Replacing this 30 amino acid peptide with a previously described sequence containing residues from the wild-type protein imbedded in a sequence matrix of mostly alanines and serines, we used a combinatorial mutagenesis and selection strategy to define residual wild-type residues that are not replaceable with alanine or serine, because they are needed for specific recognition of methionine tRNA. Four were identified, of which three have functional side chains (Asn, Arg, Lys). These four and a fifth (Trp) that was previously identified are located at the end of the helix and within the loop, lie on the same side of the structure, and span a distance of about 20 A. We conclude that, within the alanine, serine sequence matrix, only a few non-alanine, non-serine residues in the specificity-determining part of the structure are essential.

Acylation↗
PubMed1994

A phase I trial of intraperitoneal carboplatin and etoposide with granulocyte macrophage colony stimulating factor support in patients with intraabdominal malignancies.

BACKGROUND: Although somewhat controversial, there are data to suggest that patients with ovarian cancer may experience a survival advantage if the dose intensity of platinum-containing regimens can be maximized. Administration of chemotherapeutic agents via the intraperitoneal route offers the opportunity to increase dose intensity of several chemotherapeutic agents. METHODS: The authors conducted a Phase I trial of intraperitoneal carboplatin and etoposide in combination with granulocyte macrophage colony stimulating factor (GM-CSF) in an attempt to determine the maximum tolerated dose of carboplatin. The starting dose for carboplatin was 300 mg/m2 and for etoposide 400 mg/m2. The dose of carboplatin was escalated while the etoposide was maintained at the initial dose. The total dose of each agent was calculated and given daily over 3 days in amounts equal to one-third of the total dose. On day 1 of therapy, one-third of the dose was mixed in 2 liters of dextrose (D5W) and administered intraperitoneally (IP) as rapidly as possible. On Days 2 and 3, one-third of the dose was mixed in 1 liter of D5W and administered similarly. GM-CSF was begun on Day 4 as a subcutaneous injection at a dose of 500 micrograms/m2/d. RESULTS: Unacceptable hematologic toxicity was encountered at a carboplatin dose of 800 mg/m2; therefore, a carboplatin dose of 600 mg/m2 is recommended for Phase II studies. An overall response rate of 54% with a complete response rate of 17% was observed in patients with ovarian cancer. The overall response rate for all patients was 45%. CONCLUSION: Because of the significant toxicity encountered in this study, it is recommended that this regimen be used only in the context of a clinical study. The recommended Phase II study dose for this combination is carboplatin 600 mg/M2 and a total dose of etoposide 400 mg/M2 total dose given as three equal parts IP over 3 days. GM-CSF should begin on Day 4 at a dose of 500 micrograms/m2/day subcutaneously and should continue until the absolute neutrophil count is greater than 1000 granulocytes on 3 successive days.

Abdominal Neoplasms↗
PubMed1994

Purification and characterization of S-adenosylmethionine-protein-arginine N-methyltransferase from rat liver.

A protein methylase I (S-adenosylmethionine-protein-arginine N-methyltransferase; EC 2.1.1.23), with a high specificity for recombinant heterogeneous nuclear ribonucleoprotein particle (hnRNP) protein A1, was purified from rat liver. The purification method is simple and rapid; a single initial step of DEAE-cellulose DE-52 chromatography resulted in a 114-fold enrichment from the cytosol, and subsequent Sephadex G-200 chromatography and f.p.l.c. yielded a homogeneous preparation. Ouchterlony double-immunodiffusion analysis indicated that the rat liver enzyme is immunologically different from an analogous enzyme from the calf brain, nuclear protein/histone-specific protein methylase I [Ghosh, Paik and Kim (1988) J. Biol. Chem. 263, 19024-19033; Rajpurohit, Lee, Park, Paik and Kim (1994) J. Biol. Chem. 269, 1075-1082]. The purified enzyme has a molecular mass of 450 kDa on Superose chromatography and 110 kDa on SDS/PAGE, indicating that it is composed of four identical-size subunits. The Km values for protein A1 and S-adenosyl-L-methionine were 0.54 x 10(-6) and 6.3 x 10(-6) M respectively. S-Adenosyl-L-homocysteine and sinefungin were effective inhibitors of the enzyme with Ki values of 8.4 x 10(-6) M and 0.65 x 10(-6) M respectively. Bivalent metal ions such as Zn2+, Mn2+ and Ni2+ were particularly toxic to the enzyme; at 1 mM Zn2+, 99% of the activity was inhibited. In addition, 50% of the enzyme activity was lost by treatment with 0.12 mM p-chloromercuribenzoate, indicating a requirement for a thiol group for enzyme activity. Glycerol, a compound often used to prevent enzyme inactivation, inhibited over 80% of the activity when present in the reaction mixture at a concentration of 20%.

Animals↗
PubMed1994

Genomic organization of Borna disease virus.

Borna disease virus is a neurotropic negative-strand RNA virus that infects a wide range of vertebrate hosts, causing disturbances in movement and behavior. We have cloned and sequenced the 8910-nucleotide viral genome by using RNA from Borna disease virus particles. The viral genome has complementary 3' and 5' termini and contains antisense information for five open reading frames. Homology to Filoviridae, Paramyxoviridae, and Rhabdoviridae is found in both cistronic and extracistronic regions. Northern analysis indicates that the virus transcribes mono- and polycistronic RNAs and uses termination/polyadenylylation signals reminiscent of those observed in other negative-strand RNA viruses. Borna disease virus is likely to represent a previously unrecognized genus, bornaviruses, or family, Bornaviridae, within the order Mononegavirales.

Amino Acid Sequence↗
PubMed1994

Characterization of a novel intracellular sphingolipid-gated Ca(2+)-permeable channel from rat basophilic leukemia cells.

Sphingolipids stimulate the release of Ca2+ from intracellular stores. However, the mechanism by which this process occurs has not been characterized. Through single-channel recording from microsomes incorporated into planar lipid bilayers, we describe a novel channel that gates Ba2+ in response to sphingosylphosphorylcholine. The channel is both ligand-gated and voltage-modulated. Maximal open probability is observed between -10 and -20 mV and has a relatively high conductance (160 picosiemens with 53 mM Ba2+). We also observe that Ca2+ efflux from permeabilized rat basophilic leukemia cells is not antagonized by heparin, La3+, Ni2+, nifedipine, or omega-conotoxin GVIa. The sphingolipid-gated Ca(2+)-permeable channel is therefore a new member of the Ca(2+)-permeable, ligand-gated channel family.

Animals↗
PubMed1994

Hydrodynamic steering effects in protein association.

Protein-ligand reaction rates are often limited by the rate of diffusional encounter of the protein and ligand in solution. Reaction rates, however, can be much greater than expected, given the necessity for correct orientation before reaction. A number of forces can affect the orientation of the protein and ligand in solution, and thus increase the reaction rate. We have considered hydrodynamic forces, produced when water molecules between protein and ligand must be pushed out of the way to allow their encounter. We have used the cleft enzymes as a model system, as they could be expected to show strong hydrodynamic effects. One particular type of hydrodynamic interaction stands out: a steering torque which occurs when the enzyme and substrate move toward each other in solution. The magnitude of this steering torque is compared to the mutual torque experienced by interacting "protein-sized" dipoles in solution. A simple model is used to demonstrate that the hydrodynamic steering torque can be 2 orders of magnitude greater than the electrostatic torque.

Biophysical Phenomena↗
PubMed1994

Purification and characterization of nitrogenase from a delta nifW strain of Azotobacter vinelandii.

Deletion of the nifW gene in Azotobacter vinelandii yields a strain (DJ224) that grows poorly under N2 fixing conditions (Jacobson, M. R., Cash, V. L., Weiss, M. C., Laird, N. F., Newton, W. E., and Dean, D. R. (1989) Mol. & Gen. Genet. 219, 49-57). Here we report the purification of nitrogenase from DJ224. The purified Fe protein was indistinguishable from wild-type. The MoFe protein was indistinguishable from the wild-type MoFe protein by the criteria of sodium dodecyl sulfate-polyacrylamide gel electrophoresis, native gel electrophoresis, two-dimensional gel electrophoresis, metal analysis, UV/visible and EPR spectroscopies. It was different by the criteria of CD spectroscopy and specific activities. At a 5:1 molar ratio of Fe protein to MoFe protein, H2 evolution under argon was identical to wild-type, C2H2 reduction was inhibited by 27%, N2 reduction was inhibited by 38%, and CO inhibited H2 evolution by 17%. The above data show that the nifW gene product is not required for: 1) detectable alteration of the polypeptide; 2) the synthesis of the metal portion of FeMo cofactor; or 3) FeMo cofactor insertion. The MoFe protein synthesized in the absence of NifW appears to have an alteration near the FeMo cofactor site, possibly at homocitrate, which causes differential inhibition of different substrates.

Amino Acid Sequence↗
PubMed1994

Enzymatic methylation of recombinant heterogeneous nuclear RNP protein A1. Dual substrate specificity for S-adenosylmethionine:histone-arginine N-methyltransferase.

We have previously reported the existence of two different molecular species of protein methylase I (S-adenosylmethionine:protein-arginine N-methyltransferase, E.C. 2.1.1.23) in calf brain, one specific for myelin basic protein and the other for histone (Ghosh, S. K., Paik, W. K., and Kim, S. (1988) J. Biol. Chem. 263, 19024-19033). In the present study, however, we report that heterogeneous ribonucleoprotein particle protein A1 is most likely an in vivo substrate for the "histone-specific protein methylase I." The unmethylated recombinant protein A1 has been found to be a much superior methyl acceptor for the enzyme than histone with a Km value two orders of magnitude lower (0.19 microM) than that for histone (21 microM). Myelin basic protein, a specific inhibitor for histone protein methylase I, exhibited a lower IC50 for protein A1 methylation (IC50 = 33 microM) compared with histone methylation (IC50 = 220 microM) and competitively inhibited the former with a Ki value of 1.3 x 10(-6) M. The extent of inhibition of protein A1 and histone methylation by the polyclonal antibodies prepared against purified "histone protein methylase I" was identical. Maximally, 1.08-mol methyl groups were incorporated per mol of protein A1, which was 27-fold higher than that of histone (0.04 mol/mol of histone). HPLC analysis of the enzymatically methylated amino acid residues in protein A1 revealed the formation of NG-monomethylarginine and NG,NG-dimethylarginine. The ratio of NG,NG-dimethylarginine/NG-monomethylarginine increased as a function of incubation period; however, NG,N'G-dimethylarginine was not detectable. Proteolytic cleavage of the methyl-3H-labeled recombinant protein A1 by trypsin and Staphylococcus aureus V8 endoprotease indicated that protein A1 possesses multiple sites for methylation, one of which was identified as residue 194 arginine, which coincided with the in vivo methylation site.

Amino Acid Sequence↗
PubMed1994

Sustained-release methotrexate for intracavitary chemotherapy.

Methotrexate was encapsulated into a lipid-based drug-delivery system to create a slow-release formulation (Depo/methotrexate) for intracavitary administration. Depo/methotrexate was stable in storage at 4 degrees C for > 4 months. In human plasma at 37 degrees C, the half-life of drug release was 40 days. After intraperitoneal injection of Depo/methotrexate in mice, the intraperitoneal apparent half-life of free methotrexate was 39.6 h, in contrast to a half-life of 0.5 h for the unencapsulated standard methotrexate (std/methotrexate). In L1210 murine leukemia model, the potency of a single dose of Depo/methotrexate was 334-fold higher, the increased life-span (ILS) was 2-fold greater, and the therapeutic index was 2-fold higher than a single dose of std/methotrexate.

Animals↗
PubMed1994

Crystallization and preliminary X-ray crystallographic analysis of phospholipid transfer protein from maize seedlings.

Phospholipid transfer protein from maize seedlings has been crystallized using trisodium citrate as precipitant. The crystal belongs to the orthorhombic space group P2(1)2(1)2(1) with unit cell dimensions of a = 24.46 A, b = 49.97 A, and c = 69.99 A. The presence of one molecule in the asymmetric unit gives a crystal volume per protein mass (Vm) of 2.36 A3/Da and a solvent content of 48% by volume. The X-ray diffraction pattern extends at least to 1.6 A Bragg spacing when exposed to both CuK alpha and synchrotoron X-rays. A set of X-ray data to approximately 1.9 A Bragg spacing has been collected from a native crystal.

Amino Acid Sequence↗