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K Matsubara

Publications and source records attributed to K Matsubara.

At least 523 records · Page 29Linked to original sources
PubMed1982

Replication of lambda dv plasmid in vitro promoted by purified lambda O and P proteins.

An in vitro system for replication of lambda dv plasmid DNA has been constructed. This system consists of an ammonium sulfate fraction from Escherichia coli extract, exogenously added purified lambda O and P proteins, and lambda dv DNA in closed circular form. More than 85% of the added template DNA replicated semiconservatively. In the same system, another plasmid, pBR322, also replicated, but less efficiently than lambda dv. Furthermore, its replication was independent of O and P proteins. Inhibitors of DNA gyrase entirely blocked the replication activity, whereas rifampicin, an inhibitor of RNA polymerase, showed a significant effect only when added prior to initiation of the DNA replication. DNA replication was initiated from a region near to or within the four direct repeats in lambda origin (lambda ori) and proceeded bidirectionally, as examined by DNA chain elongation termination with dideoxy CTP. A cloned DNA carrying a 350-base-pair region including the initiation site also initiated replication, dependent on O and P proteins, and its initiation occurred at the same position as with native lambda dv DNA. An A + T-rich structure neighboring the repeats was found to be essential for lambda DNA replication. Regions corresponding to ice and oop were not required for O,P-dependent initiation.

Bacteriophage lambda↗
PubMed1982

Acute effects of ethanol and acetaldehyde on plasma phosphate level.

Oral administration of ethanol in a dose of 65 mmol kg-1 produced marked change of plasma phosphate level in rabbits. Hypophosphataemia was observed for the first 2 h after administration followed by significant increase of plasma phosphate at 5 h. Hypophosphataemia did not appear when ethanol was given to the rabbits pretreated with pyrazole. When animals were injected with disulfiram in advance, the duration of hyperphosphataemia due to ethanol was prolonged. Administration of acetaldehyde at a dose of 1.5 mmol kg-1 produced hyperphosphataemia. In this study, plasma phosphate was not associated with change in calcium level. These results suggest that the hypophosphataemia observed was related to the metabolic process of ethanol utilizing alcohol dehydrogenase, and that acetaldehyde, a metabolite of ethanol, might induce the hyperphosphataemia in the animals.

Acetaldehyde↗
PubMed1981

Structure of the baboon endogenous virus genome: cloning of circular virus DNA in bacteriophage lambda.

Linear, small and large circular forms of unintegrated viral DNAs were detected in Hirt supernatant fraction of human cultured cells infected with baboon endogenous virus M7. The circular M7 DNAs were cloned in bacteriophage lambda, Charon 28. Seventeen independent clones were isolated and analyzed by restriction endonuclease mapping. Nine clones were carrying a viral sequence of 8.6 kilobase pairs (kb) with two tandem repeats of 0.6 kb, which correspond to the large circular form of the unintegrated M7 DNA. Eight other clones had the viral insert of 8.0 kb, i. e., the small circular form, and were deleted one of the repeated sequences. The repeated sequences correspond to the long terminal repeats of 0.6 kb, located at both ends of the linear M7 DNA of 8.6 kb. One of the recombinants of the large circular M7 DNA had an inversion of 2.5 kb. One end of the inverted sequence was near the terminus of the long terminal repeats and the other in the gag gene region. The inversion seems to be occurred by integration of a viral DNA within itself during early periods of infection. The mechanism of the processes leading to integration is discussed from the structure of these unintegrated M7 DNAs as the precursors.

Bacteriophage lambda↗
PubMed1981

Purified bacteriophage lambda O protein binds to four repeating sequences at the lambda replication origin.

The bacteriophage lambda O protein is needed for initiation of lambda DNA replication. Several lines of evidence suggest that initiation requires that this protein interacts with a specific sequence called ori (for origin) in lambda DNA. We have purified this protein to near homogeneity and studied the protection against nuclease cleavage of the origin DNA sequences. Our data demonstrate that the O protein binds within an interval of about 95 base pairs (bp), which contains four tandemly arranged 19bp repeating sequences, ATCCCTCAAAACGA (G)GG GAT(A). At a low concentration of O protein, the inner two repeats are primarily covered, while binding to the outer two repeats requires a high concentration of O protein. From the molecular size of O protein (32,000 daltons), and the internal symmetry in each 19bp repeat, we inferred that the O protein may bind in dimeric form, and that the 95bp region may be filled only when four such dimers have bound. This interaction is discussed in connection with the "activation" of the ori by O protein leading to initiation of DNA synthesis.

Bacteriophage lambda↗
PubMed1981

Purification of bacteriophage lambda O protein that specifically binds to the origin of replication.

By means of a nitrocellulose filter binding assay, DNA binding activities among proteins fractionated from extracts of Escherichia coli carrying lambda dv have been surveyed. An activity was found that binds specifically to a fragment of 164 base pairs that specifies the lambda replication origin (lambda ori). This activity was not detected in an extract of cells not carrying the lambda dv plasmid. The activity was detected in extracts of cells carrying a hybrid plasmid in which the entire lambda O gene had been cloned and placed under the control of the lac promoter. Deletion of a 60 base pair segment in the 'amino-terminal region' of the O gene abolished this activity, indicating that the lambda ori binding protein is coded for by the lambda O gene. The ori-specific binding protein was purified by five fractionation steps. The most purified preparation consists of a major polypepide that migrates with a molecular weight of 32,000 in SDS-polyacrylamide gel electrophoresis. Binding of O protein to ori occurs in the absence of other protein aceous components.

Bacteriophage lambda↗
PubMed1981

Nine unique repeating sequences in a region essential for replication and incompatibility of the mini-F plasmid.

The nucleotide sequence of a 2248 bp portion of the plasmid mini-F has been determined. This region includes the replication origin and all of the plasmid-coded information required for replication. The same region is also capable of expressing incompatibility. A striking feature of the sequence is the presence of nine 19-bp repeating units. Four of these repeats, all arranged in one direction, comprise a cluster, and the remaining five, all arranged in the opposite direction, comprise another cluster. These clusters are separated by a region of about 850 bp that encodes a hypothetical 29-kd polypeptide. This region has sequences highly homologous to those found in the origin regions of the Escherichia coli (Sugimoto et al., 1979; Meijer et al., 1979) and Salmonella typhimurium (Zyskind and Smith, 1980) genomes.

Cloning, Molecular↗
PubMed1981

Replication control and switch-off function as observed with a mini-F factor plasmid.

Mini-F is a fragment of the F plasmid, consisting of 9,000 base pairs, which carries all of the genes and sites required for replicon maintenance and control. Its copy number is one to two per chromosome. This plasmid is joined to ColE1, whose copy number is 16 to 20. Under normal circumstances the composite plasmid replication exhibited ColE1 characteristics, maintaining a high copy number. However, when ColE1 replication was inhibited by deoxyribonucleic acid polymerase I inactivation, its replication exhibited mini-F characteristics, maintaining a low copy number. These observations are in complete agreement with those of Timmis et al. (Proc. Natl. Acad. Sci. U.S.A. 71:4556-4560, 1974), who examined the behavior of a recombinant plasmid formed between pSC101 and ColE1. The transition from high to low copy number allowed us to examine the control system acting in cells carrying plasmids exhibiting intermediate copy numbers. The initiation of the mini-F replication system as represented by deoxyribonucleic acid synthesis of the composite plasmid was completely blocked when there were multiple copies of mini-F in a cell. It was not restored until the copy number was lowered to one to two, after which replication was first detected. ppF, a mini-F replicon packaged in a phage lambda head behaved similarly: its replication was completely shut off when the resident mini-F genome copy number was high and was inhibited partially when the resident mini-F genome copy number was low. These experiments clearly demonstrate that there is a switch-off mechanism acting on deoxyribonucleic acid synthesis (initiation) in a cell carrying mini-F, and its intensity is related to the plasmid copy number. This result supports the "inhibitor dilution model" proposed by Pritchard et al. (Symp. Soc. Gen. Microbiol. 19:263-297, 1969). The nature of the hypothetical inhibitor is discussed.

DNA Replication↗
PubMed1981

Pharmacology of 2-o-chlororbenzoyl-4-chloro-N-methyl-N alpha-glycylglycinanilide hydrate (45-0088-S), a compound with benzodiazepine-like properties.

2-o-Chlorobenzoyl-4-chloro-N-methyl-N alpha-glycylglycinanilide hydrate (45-0088-S), a dipeptido-aminobenzophenone compound, inhibited pentetrazole-induced convulsion and suppressed electroshock-induced fighting behavior in mice, reduced conflict behavior in rats and monkeys, and induced muscle relaxation in cats at lower doses than did diazepam. This pharmacological profile suggests that 45-0088-S can be an active, water-soluble antianxiety agent in man.

Aggression↗