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Biomedical subjects

S Kawabata

Publications and source records attributed to S Kawabata.

At least 217 records · Page 12Linked to original sources
PubMed1988

A mutant rat strain deficient in induction of a phenobarbital-inducible form of cytochrome P-450 in liver microsomes.

Two phenobarbital-inducible forms of cytochrome P-450, P-450(PB-1), and P-450(PB-4), were purified from the liver microsomes of phenobarbital-treated rats and identified with P-450b and P-450e, respectively. It was found, however, that the content of P-450(PB-4) in the liver microsomes of a strain of SD rat, Qdj:SD, was very low even after phenobarbital-induction. The levels of the mRNAs for P-450(PB-1) and P-450(PB-4) were separately determined using Northern blot hybridization with specific oligonucleotide probes. It was found that the level of P-450(PB-4) mRNA in the livers of phenobarbital-treated Qdj:SD rats was much lower than that of phenobarbital-treated Slc:SD rats. Slc:SD rats are widely used in Japanese laboratories. Genetic analysis using the crossbred animals between Qdj:SD and Slc:SD rats showed that the low expression of P-450(PB-4) in Qdj:SD rats is a recessive trait and is caused by a single gene mutation. However, no difference in the 5' flanking region in P-450(PB-4) gene was found between Qdj:SD rats and Slc:SD rats.

Animals↗
PubMed1988

Amino acid sequence of a coagulant enzyme, flavoxobin, from Trimeresurus flavoviridis venom.

The amino acid sequence of a coagulant enzyme, named flavoxobin, isolated from the venom of Trimeresurus flavoviridis (the habu snake) was determined by sequencing the S-pyridylethylated derivative of the protein and its peptides generated by chemical (cyanogen bromide and hydroxylamine) and enzymatic (clostripain, Staphylococcus aureus V8 protease, Achromobacter protease I, and elastase) cleavages. Hydrazinolysis was also employed to determine the C-terminal amino acid. The enzyme consisted of 236 amino acids and had a calculated molecular weight of 25,744. Flavoxobin was found to be highly (69%) homologous in sequence to batroxobin, a coagulant enzyme from the venom of Bothrops atrox, and 27, 39, and 31% homologous to bovine thrombin, bovine trypsin, and human kallikrein, respectively. The sequence around the active site serine residue deduced from the homology relationship was Phe-Asp-Ser-Gly-Thr, which is different from the common sequence, Gly-Asp-Ser-Gly-Gly, for most serine proteases. Flavoxobin appears to be similar in secondary structure composition to batroxobin.

Amino Acid Sequence↗
PubMed1988

A characteristic property of vitamin K-dependent plasma protein Z.

Evidence is presented for rapid, limited proteolysis of protein Z by alpha-thrombin. This alpha-thrombin-catalyzed proteolysis of protein Z occurred at a single peptide linkage, between Arg-365 and Gly-366, located in the COOH-terminal portion. The resulting NH2-terminal large fragment (PZt) and the COOH-terminal peptide (C-peptide) were isolated and chemically characterized. The C-peptide consisted of 31 amino acid residues including one galactosamine-type Thr residue and was assigned to the position from Gly-366 to the COOH-terminal residue of Val-396 in protein Z. The NH2-terminal large fragment, PZt, constituted the remainder of protein Z. The abilities to bind calcium of intact protein Z, PZt, and the derivative of protein Z devoid of the NH2-terminal gamma-carboxyglutamic acid (Gla) domain (Gla-domainless), prepared with the known chymotrypsin treatment, were examined by equilibrium dialysis. The results indicated that intact protein Z and PZt contain four calcium binding sites with dissociation constants of 0.1 mM. Moreover, the Scatchard plot analysis showed positive cooperativity, suggesting the presence of at least two initial sites for calcium binding. In contrast, the Gla-domainless protein Z had no calcium binding site, indicating that the domain of protein Z functional for calcium binding occurs within the NH2-terminal Gla domain. This differed from factor X, factor IX, protein S, and protein C, all of which contain one or two calcium binding site(s) independent on their Gla-domains.

Amino Acid Sequence↗
PubMed1988

Molecular cloning and nucleotide sequence of cDNA of microsomal carboxyesterase E1 of rat liver.

cDNA clones of the mRNA for rat liver carboxyesterase E1, one of the carboxyesterases exclusively located on the luminal side of microsomal vesicles, were isolated. Sequence analysis of 2 kbp long cDNA revealed the primary structure of carboxyesterase E1, which consisted of 549 amino acids (Mr 60, 171.71) and contained an extra peptide of 18 amino acids at the NH2-terminus of the mature enzyme. Comparison of the deduced primary structure and sequences of some proteolytic fragments of the purified enzyme indicated the multiplicity of the enzyme. The extra peptide at the NH2-terminal had features in common with the signal peptides of most secretory proteins. However, no polar amino acid residues existed before the hydrophobic core of the signal peptide. A new interpretation is proposed to explain how the signal peptide without the NH2-terminal polar residues works. A tetrapeptide (KDEL) which was shown to keep a few microsomal proteins in the lumen of the endoplasmic reticulum was not found in the primary structure of carboxyesterase E1, which suggested the existence of another mechanism for retention of proteins in the lumen of endoplasmic reticulum. Carboxyesterase E1 showed significant homology with the COOH-terminal portion of thyroglobulin.

Amino Acid Sequence↗
PubMed1988

A new trisaccharide sugar chain linked to a serine residue in bovine blood coagulation factors VII and IX.

A new trisaccharide sugar chain was identified in bovine blood coagulation factors VII and IX. A pentapeptide isolated from factor VII contained Ser-52, which could not be identified with a gas-phase sequencer, suggesting an unknown substituent on the serine residue (Takeya, H. et al. (1988) J. Biol. Chem., in press). The same results were obtained for a pentapeptide containing Ser-53 of factor IX. Component sugar analysis revealed that the peptide contained 1 mol of glucose and 2 mol of xylose. This sugar component was also confirmed by high-resolution fast atom bombardment mass spectrometric analysis of the pentapeptide. The trisaccharide was released from the peptides by means of beta-elimination reaction and its reducing end was coupled with 2-aminopyridine. The fluorescent pyridylamino (PA-) derivative of the trisaccharide was purified by gel-filtration and reversed-phase HPLC. The sugar composition of the PA-trisaccharide was found to be 2 mol of xylose and 1 mol of PA-glucose. These results indicate the existence of a (Xyl2)Glc-Ser structure in factors VII and IX.

Amino Acids↗
PubMed1988

Blood clotting factor IX Niigata: substitution of alanine-390 by valine in the catalytic domain.

Factor IX Niigata is a mutant factor IX responsible for the moderately severe hemophilia B in a patient who has a normal level of factor IX antigen with reduced clotting activity (1-4% of normal). We reported previously that the purified mutant protein could be converted to the factor IXa beta form by factor XIa/Ca2+ at a rate similar to that in the case of normal factor IX, but the resulting mutant factor IXa beta could not activate factor X in the presence of factor VIII, Ca2+, and phospholipids (Yoshioka, A. et al. (1986) Thromb. Res. 42, 595-604). In the present study, we analyzed factor IX Niigata at the structural level to elucidate the molecular abnormality responsible for the loss of clotting activity. Amino acid sequence analysis of a peptide obtained on lysyl endopeptidase digestion, coupled with subsequent SP-V8 digestion, demonstrated that the alanine at position 390 was substituted by valine in the catalytic domain of the factor IX Niigata molecule.

Alanine↗
PubMed1988

Cloning and sequence of the gene encoding the major structural component of mannose-resistant fimbriae of Serratia marcescens.

Serratia marcescens US46, a human urinary tract isolate, exhibits mannose-resistant hemagglutination and agglutinates yeast cells, thereby indicating that it has two types of adhesins. We constructed a cosmid library for the DNA of this organism and isolated DNA clones carrying genes for mannose-sensitive (MS) and mannose-resistant (MR) fimbriae. On introduction of the cloned genes into Escherichia coli K-12, MS and MR fimbriae were formed. These fimbriae were functionally and morphologically indistinguishable from those of S. marcescens. Subcloning of these gene clusters revealed that the genes encoding MS fimbriae reside on a 9-kilobase (kb) DNA fragment, while those encoding MR fimbriae are present on a 12-kb fragment. Transposon insertion and maxicell analyses revealed that formation of MR fimbriae is controlled by several genes which reside on the 9-kb fragment. The nucleotide sequence of smfA, the gene encoding the major structural component of MR fimbriae, revealed that this gene encodes a 174-amino-acid polypeptide with a typical procaryotic signal peptide. The primary structure of the smfA product showed significant homology with the primary structure of the E. coli fimbrial subunit.

Amino Acid Sequence↗
PubMed1988

Changes of portal systemic shunting after sclerotherapy and/or partial splenic embolization observed by thallium-201 per rectal administration.

Tl-201 per-rectal scintigraphy was performed before and after injection sclerotherapy of esophageal varices and/or partial splenic embolization in patients with esophageal varices to observe the changes of portal systemic shunting by these treatments. The changes in the heart-to-liver uptake ratio of Tl-201 after injection sclerotherapy ranged widely from a decrease to an increase with a mean change rate (mean of the absolute value of change rate) and one standard deviation of 19.0 +/- 19.9%. By partial splenic embolization the heart-to-liver uptake ratio was decreased significantly with the mean change rate and one standard deviation of 26.2 +/- 17.4% (P less than .05). This noninvasive and simple method seems to be useful in evaluating pathophysiological changes of portal systemic shunting observed from inferior mesenteric vein blood flow after treatments of portal circulation system.

Administration, Rectal↗
PubMed1988

Changes of portal systemic shunting after sclerotherapy and/or partial splenic embolization observed by Tl-201 per-rectal administration.

Tl-201 per-rectal scintigraphy was performed before and after injection sclerotherapy of esophageal varices and/or partial splenic embolization in patients with esophageal varices to observe the changes of portal systemic shunting by these treatments. The changes in the heart-to-liver uptake ratio of Tl-201 after injection sclerotherapy ranged widely from a decrease to an increase with the mean change rate (mean of the absolute value of change rate) +/- 1 SD of 19.0% +/- 19.9%. By partial splenic embolization the heart-to-live uptake ratio was decreased significantly with the mean change rate +/- 1 SD of 26.2% +/- 17.4% (P less than 0.05). This noninvasive and simple method seems to be useful in evaluating pathophysiological changes of portal systemic shunting observed from inferior mesenteric vein blood flow after treatments of portal circulation system.

Administration, Rectal↗
PubMed1987

Structural comparison of bovine erythrocyte, brain, and liver NADH-cytochrome b5 reductase by HPLC mapping.

NADH-cytochrome b5 reductases purified from bovine erythrocytes and from bovine brain and liver microsomes solubilized with lysosomal protease were subjected to structural analysis by using HPLC mapping, amino acid analysis of the resulting peptides, and NH2-terminal sequence analysis of apoproteins. HPLC maps of the tryptic peptides derived from these enzymes were very similar to each other, and amino acid analysis of the HPLC-separated peptides indicated that the structures of these enzymes are identical except for the NH2-terminal region. The NH2-terminal sequence of the brain enzyme determined by automated Edman degradation was as follows: NH2-Phe-Gln-Arg-Ser-Thr-Pro-Ala-Ile-Thr-Leu-Glu-Asn-Pro-Asp- Ile-Lys-Tyr-Pro-Leu-Arg-Leu-Ile-Asp-Lys-Glu-Val-Ile- This sequence is identical to that of liver enzyme except that the liver enzyme started at the 3rd Arg or 4th Ser. The NH2-terminal amino acid residue of the soluble erythrocyte enzyme was not detected by automated Edman degradation. The sequence analysis of a tryptic peptide from the erythrocyte enzyme indicated that Leu is present before the NH2-terminal Phe of the brain enzyme. The recently reported sequence of the apparently identical protein (Ozols et al. (1985) J. Biol. Chem. 260, 11953-11961) differs in two amino acid assignments from our sequence.

Amino Acid Sequence↗
PubMed1987

Purification and amino acid sequence of Kunitz-type protease inhibitor found in the hemocytes of horseshoe crab (Tachypleus tridentatus).

A low molecular weight protein protease inhibitor was purified from Japanese horseshoe crab (Tachypleus tridentatus) hemocytes. It consisted of a single polypeptide with a total of 61 amino acid residues. This protease inhibitor inhibited stoichiometrically the amidase activity of trypsin (Ki = 4.60 X 10(-10) M), and also had inhibitory effects on alpha-chymotrypsin (Ki = 5.54 X 10(-9) M), elastase (Ki = 7.20 X 10(-8) M), plasmin, and plasma kallikrein. However, it had no effect on T. tridentatus clotting enzyme and factor C, mammalian blood coagulation factors (activated protein C, factor Xa and alpha-thrombin), papain, and thermolysin. The complete amino acid sequence of this inhibitor was determined and its sequence was compared with those of bovine pancreatic trypsin inhibitor (BPTI) and other Kunitz-type inhibitors. It was found that the amino acid sequence of this inhibitor has a high homology of 47 and 43% with those of sea anemone inhibitor 5-II and BPTI, respectively. Thus, this protease inhibitor appeared to be one of the typical Kunitz-type protease inhibitors.

Amino Acid Sequence↗
PubMed1987

Nucleotide sequence of the staphylocoagulase gene: its unique COOH-terminal 8 tandem repeats.

The entire staphylocoagulase gene of Staphylococcus aureus strain BB was cloned on a MboI restriction endonuclease fragment inserted into pAT153 plasmid vector. The staphylocoagulase was expressed in Escherichia coli, as judged by the formation of a fibrin halo on an agar plate containing rabbit plasma and bovine fibrinogen. We have determined the complete nucleotide sequence of the staphylocoagulase gene by the dideoxynucleotide chain termination method. The deduced amino acid sequence consisted of 715 residues including a signal peptide of 26 residues. Therefore, the predicted molecular weight of the mature protein was 77,337. This sequence was corroborated by reference to the amino acid compositions of 30 lysyl endopeptidase peptides and the sequences of 12 of these peptides isolated from the purified staphylocoagulase. The 5'-flanking region was found to contain a putative Shine-Dalgarno sequence and a putative "-10" element for transcription. The COOH-terminal stretch of 216 amino acids of staphylocoagulase was composed of 8 tandem repeats each consisting of 27 amino acid residues. The amino acid sequence of staphylocoagulase derived from strain BB showed 57% identity with that of the chymotryptic 43-kDa fragment of staphylocoagulase isolated previously from strain 213 (Kawabata, S., Miyata, To., Morita, T., Miyata, Ta., Iwanaga, S., & Igarashi, H. (1986) J. Biol. Chem. 261, 527-531).

Amino Acid Sequence↗
PubMed1987

Antigenic determinants on fimbriae of Serratia marcescens US5 analyzed using monoclonal antibodies.

The antigenic sites on small thin fimbriae of Serratia marcescens strain US5 were investigated using immunoelectron microscopy and monoclonal antibodies (MAbs). Negative staining of the fimbriae after treatment with MAbs showed a regularly spaced arrangement of the antibody molecules. When the subunit peptide was subjected to immunoblotting using the MAbs, a single band with a molecular weight of approximately 19kD was evident. This binding of the MAbs to the subunit peptide was completely abrogated after treatment with 2-mercaptoethanol, thereby suggesting the important role of disulfide linkage in the maintenance of the conformation of the antigenic site reacted with MAbs. Amino acid analysis of the subunit peptide revealed two cysteine residues, and cysteine residues were absent in the N-terminal portion.

Amino Acid Sequence↗