Conversion of a plasma group-specific component (vitamin D binding protein) to a macromolecule by a protein factor in tissue extracts.
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Actins↗
Biomedical subjects
T Shinomiya
Publications and source records attributed to T Shinomiya.
Characterization of antibody to 2'-5' linked oligoadenylic acid.
Antibody to 2'-5' linked triadenylate, A2'p5'A2'p5'A, was prepared by injecting bovine serum albumin conjugated with A2'p5'A2'p5'A into rabbits. The [8-14C]-A2'p5'A2'p5'A used in the study was synthesized by nonenzymatic template-directed condensation of [8-14C]AMP and 5'-phosphorimidazolide of A2'p5'A on poly U. The specificity of the antibody was determined by the inhibition of the binding of the radioisotope probe to the antibody by a competitor using the ammonium sulfate precipitation method. The cross reactivity of a number of modified oligonucleotides has been evaluated in order to gain information on the structural feature involved in the binding to the antibody. The adenine moiety and 2'-5' internucleotide linkage are crucial regions for the binding. The antibody reacted not only on A2'p5'A2'p5'A and A2'p5'A2'p5'A2'p5'A, but also on a cordycepin analogue of the trimer, (3'dA)2'p5'(3'dA)2'p5'(3'dA), significantly. Considerable cross reaction was observed with oligoadenylates bearing both 2'-5' and 3'-5' internucleotide linkages, while there was little cross reaction with 3'-5' linked oligoadenylates. The antibody showed very low or no affinity for adenosine, adenine mononucleotides, other nucleotides and other 2'-5' linked trinucleotides. The cross reactivity of the modified oligonucleotides is indicated to have relation to the biological activity of the corresponding 5'-phosphorylated oligonucleotides, so far reported. The antibody was further purified by affinity chromatography using RNA-Sepharose and AMP-Sepharose giving high specificity.
Adenine Nucleotides↗
A new aspect of a restriction endonuclease Tth111 I. It has a degenerated specificity (Tth111 I).
We previously reported that Thermus thermophilus 111 contained two restriction enzymes, Tth111 I and Tth111 II. The former does not cleave phi X174RFDNA and the latter does. We have now found another endonuclease activity able to cleave phi X174RFDNA in the cell extract of T. thermophilus 111. The protein with this activity was purified in a homogeneous state by chromatography on cellulose phosphate, heparin-Sepharose 4B and hydroxylapatite, successively. However, this endonuclease activity was always accompanied with Tth111 I activity during the purification procedure and the purified protein also showed a strong Tth111 I activity, suggesting that the Tth111 I activity and the phi X174RFDNA-cleaving activity reside in a single molecule. The phi X174RFDNA-cleaving activity was enhanced more strongly with Mn2+ than with Mg2+ and seemed to be attributable to a relaxed specificity of Tth111 I activity as seen in the cases of EcoRI* and BamHI* Thus we designated the phi X174RFDNA-cleaving activity Tth111 I*. The molecular weight of the protein with both Tth111 I and Tth111 I* activities was determined to be about 76,000 by gel filtration on a Sephadex G-100 column and 39,000 by SDS-polyacrylamide gel electrophoresis, suggesting the enzyme to be a dimer consisting of identical polypeptide chains. The phi X174RFDNA sequences surrounding Tth111 I* cuts were determined by the chain terminator method of Sanger et al. The results confirmed that Tth111 I* recognized a degenerated form of the Tth111 I recognition sequence, i.e., a sequence such that one of the specified nucleotides in the Tth111 I recognition sequence, 5'GACNNNGTC3', was substituted with N (N stands for any of A, G, C, and T), such as 5'NACNNNGTC3', 5'GACNNNNTC3', 5'GACNNNGNC, and so on (arrows indicate cleavage sites).
Bacterial Proteins↗
[Value of immunoelectrophoresis in expert evaluation of blood stains in forensic medicine. 2].
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Blood Stains↗
Preparation and binding specificity of antibody to 2'-5' linked triadenylic acid.
Antibody to 2'-5' linked triadenylic acid, A2'p5'A2'p5'A was prepared and purified by affinity chromatography. [8-14C]-A2'p5'A2'p5'A used in immunochemical studies was synthesized by nonenzymatic template-directed condensation. The specificity of the antibody was determined by the inhibition of the binding of the radioisotope probe to the antibody by a competitor. The cross reactivity of a number of modified oligonucleotides has been evaluated in order to gain information on the structural feature involved in the binding to the antibody. The adenine moiety and 2'-5' internucleotide linkage are crucial region for the binding. The cross reactivity of the modified oligonucleotides is indicated to have relation to the biological activity of the corresponding 5'-phosphorylated oligonucleotides.
Adenine Nucleotides↗
Evidence for guanylate cyclase activity associated with hemagglutinating virus of Japan (Sendai virus).
Purified virions of HVJ (Sendai virus) were found to contain a guanylate cyclase activity that converts GTP to cyclic GMP. Activities of adenylate cyclase and 5'-nucleotidase which are frequently used as marker enzymes of cell membranes were not detected in the virus. Guanylate cyclase and virion-associated activities, neuraminidase and hemagglutinin, were co-purified during a purification of virions. Guanylate cyclase activity was not detected without disruption of the virions with a detergent, Triton X-100 or Nonident P-40. Treatment of intact HVJ with a proteolytic enzyme, trypsin or chymotrypsin, destroyed both neuraminidase and hemagglutinin; however, most of the guanylate cyclase ws retained. Guanylate cyclase activity was found in fractions containing nucleocapsids after sucrose density gradient centrifugation of disrupted virions. These results indicated that the enzyme was tightly bound to cores of HVJ and, therefore, its presence could not be explained by binding of host cell enzyme to the surface of virions. Properties of the virus-derived enzyme and particulate fractions of host cell homogenates were similar. Antiserum against nucleocapsids of HVJ inhibited guanylate cyclase activity of HVJ and particulate fractions of cells such as chorioallantoic membrane and rat liver, while soluble guanylate cyclase was not inhibited by antiserum. The biological significance and origin of guanylate cyclase found in HVJ are obscure and await further study.
Electrophoresis, Polyacrylamide Gel↗
Human bones of the burial mounds in matsuyama. Report I.
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Adult↗
A second site specific endonuclease from Thermus thermophilus 111, Tth111II.
A second site specific endonuclease with novel specificity has been purified from Thermus thermophilus strain 111 and named Tth111II. The enzyme is active at temperature up to 80 degrees C and requires Mg2+ or Mn2+ for endonuclease activity. Tth111II cleaves phi X174RFDNA into 11 fragments and lambda NA into more than 25 fragments. From the 5'-terminal sequences of TthlllII fragments of phi X174RFDNA determined by the two dimensional homochromatography and the survey on nucleotide sequence of phi X174RFDNA, it was concluded that Tth111II recognizes the DNA sequence (see former index) and cleaves the sites as indicated by the arrows.
Base Sequence↗
A spite specific endonuclease from thermus thermophilus 111, Tth111I.
A site specific endonuclease with novel specificity has been isolated from Thermus thermophilus strain 111 and named Tth111I. Tth111I cleaves lambda DNA into three fragments of 23.5, 25.7 and 50.8% of the total length, and ColE1 DNA into two fragments of nearly equal length. The sequences around Tth111I cleavage sites of ColE1 and lambda DNA were determined by the Maxam and Gilbert method and the two dimensional mapping method. The results suggest that Tth111I recognizes the DNA sequence (formula: see text) and cleaves the site as indicated by the arrows. Assuming that the first T.A pair in the sequence can be replaced for any base pair, the Tth111I recognition sequence has the symmetry with the two-fold axis as most type II restriction endonucleases do.
Bacteriophage lambda↗
A DNA methylase from Thermus thermophilus HB8.
A DNA methylase was purified in a homogeneous state from a extremely thermophilic bacterium, Thermus thermophilus HB8, by chromatography on, successively, phosphocellulose, CM-cellulose, and heparin-Sepharose. The molecular weight of the enzyme was determined to be about 44,000 by gel filtration on a Sephadex G-100 column and 41,000 by SDS-poly-acrylamide gel electrophoresis, and these findings suggest a single polypeptide enzyme. The enzyme develops maximum activity around pH 7.4 and at 70 degrees C. Enzymatic activity is completely inhibited by 0.2 M NaCl or 2 mM HgCl2. The enzyme transfers methyl groups from S-adenosyl-L-methionine to a double stranded DNA. The sole product of the reaction was identified as N-6-methyl adenine after hydrolysis of the DNA with formic acid. The enzyme kinetics obey the Michaelis-Menten equation and Km values for S-adenosylmethionine and lambda phage DNA were determined to be 0.8 muM and 10 microgram/ml, respectively. The enzyme does not transfer methyl groups to TthHB8I endonuclease digested DNA as well as the host (T. thermophilus HB8) DNA. The number of methyl groups of the fully methylated phiX174 RF DNA was about twice as many as TthHB8I endonuclease sites on the DNA. The distribution of the methyl groups of phiX174 RF DNA among the HaeIII fragments was the same as that of TthHB8I endonuclease sites, suggesting that this DNA methylase is the other component of the modification-restriction system including TthHB8I endonuclease. The enzyme probably recognizes the sequence, 5'-TCGA-3', in a double stranded DNA and probably methylates adenine in the above sequence.
Bacteriophage phi X 174↗
A second site specific endonuclease from Thermus thermophilus 111,Tth111II.
A second site specific endonuclease with a novel specificity has been isolated from Thermus thermophilus strain 111 and named Tth111II. The enzyme is active at temperature up to 80 degrees C and requires Mg2+ or Mn2+ for activity. Tth111II cleaves phi X174RFDNA into 11 fragments. From the analysis of 5' terminal sequences of the phi X174RFDNA fragments produced by Tth111II action, it was concluded that Tth111II recognized the DNA sequence (See formula in text) and cleaved the sites as indicated by arrows.
Base Sequence↗
Comparative study on R-type pyocins of Pseudomonas aeruginosa.
Four R-type pyocins (R1, R2, R3, and R4) produced by different Pseudomonas aeruginosa strains were compared with respect to their structure and action methanism. It is known that these bacteriocins have structures similar to contractile bacteriophage tails and serologically cross-react with each other, but they are distinguished according to difference in range of sensitive strains. All these pyocins were shown to arrest the synthesis of protein and nucleic acid in sensitive cells, as previously reported for pyocin R1. Action of pyocin R3 was shown to require calcium ion. Although antiserum prepared against one pyocin neutralized other R-type pyocins as well as the homologous pyocin, some differences in antigen specificity were found between pyocin R1 and pyocin R2 and R3 antibody blocking assays. Electron microscopic observation of pyocin particles treated with the antiserum pre-absorbed with heterologous pyocin revealed that specific antigens were located on the distal portion of the fibers. Comparison of protein composition showed these pyocins to be composed of essentially similar subunit proteins but a subunit protein supposed to be a main constituent of the fiber was different in its molecular weight among these pyocins. These results suggest that the main structural difference of these pyocins is to be found in the fiber.
Bacteriocins↗
[A case of death by oral poisoning with sodium nitrite].
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Adult↗
Suppressor mutation in Pseudomonas aeruginosa.
Suppressor mutations were identified in Pseudomonas aeruginosa, and a comparison was made with Escherichia coli suppressor systems. A suppressor-sensitive (sus) derivative of a plasmid, RP4 trp, and several Sus mutants of IncP1 plasmid-specific phages, were isolated by using E. coli. Plasmid RP4 trp (sus) was transferred to P. aeruginosa strains carrying trp markers which did not complement RP4 trp(sus), and Trp+ variants were selected. Some, but not all such revertants, could propagate PRD1 Sus phages, and these mutants were found to be supressor positive. Plating efficiencies of various Sus phages on these strains were compared with on E. coli strains carrying known suppressor genes. The results suggested that the Pseudomonas suppressors were probably amber suppressors. In iddition, some Sus phages (PRD1sus-55, PRD1sus-56) were obtained which, although apparently of the amber type for E. coli, were able to propagate equally well on sup+ or sup strains of P. aeruginosa. On the other hand, several mutants of phage PRR1 which were suppressed in E. coli were not suppressed by the P. aeruginosa suppressor. Suppressor-sensitive mutants were also isolated with P. aeruginosa bacteriophages E79 and D3.
Bacteriophages↗
Characterization of a bacteriophage related to R-type pyocins.
A bacteriophage with a contractile tail which shows very similar features to R-type pyocins was isolated and characterized. This phage, named PS17,was purified by DEAE-cellulose chromatography and CsCl density gradient centrifugation. It was a DNA-containing phage, and the density of the purified particles in CsCl was found to be 1.468. DNA from this phage had a density of 1.720 in CsCl, indicating its guanine plus cytosine content to be 61.2%. The head was polyhedral, 69 nm in diameter, and the tail was 150 nm in length. This phage was neutralized by antiserum preparations against five R-type pyocins, and the antiserum against this phage was active in neutralizing R-type pyocins. The properties of this phage, PS17, were compared with another similar phage, PS3, which was previously reported.
Antigens, Viral↗
Bactericidal activity of the tail of Pseudomonas aeruginosa bacteriophage PS17.
The tail of bacteriophage PS17 of Pseudomonas aeruginosa was shown to be bactericidal, and its properties were compared with those of pyocin R1. Temperature-sensitive mutants were isolated from PS17, and the products at nonpermissive temperature were morphologically characterized. Bactericidal substances were found in the lysates of such mutants that were defective in the head formation but not in the tail formation. Phage tails were purified from the lysate of one such mutant, and its chemical and biological properties were studied. Isolated tails killed sensitive cells by a single-hit process and repressed the uptake of leucine in sensitive cells. These results were consistent with the previous findings on the serological and morphological relationship between PS17 and pyocin R1. However, certain differences were also shown between them in shape and protein composition.
Antigens, Viral↗
An isochizomer of TaqI from Thermus thermophilus HB8.
A site-specific endonuclease has been isolated from Thermus thermophilus HB8 and named TthHB8I. It recognizes the same sequences as TaqI from Thermus aquaticus YT-1 does. The amount of TthHB8I in the cells was comparable to that of TaqI. T. thermophilus HB8 has an advantage over T. aquaticus YT-1 for preparation of a TaqI-like enzyme since it is easier to obtain T. thermophilus HB8 cells in quantity.
Binding Sites↗
Characterization of anti-ApApA antibodies.
Antibodies to a trinucleotide, ApApA, were prepared by injecting bovine serum albumin conjugated with ApApA into rabbits. The specificities of the antibodies were determined by estimating the inhibiton of the binding of [14C]ApApA to the antibodies by various nonradioactive mono-, oligo-, and polynucleotides, using the ammonium sulfate precipitation method. The antibodies were found to react with ApApN sequences and oligoadenylic acids, but also reacted slightly with polyadenylic acid, RNA and DNA. Significant crossreactions were observed with other oligonucleotides containing adenosine.
Adenine Nucleotides↗