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Biomedical subjects

M Sekiguchi

Publications and source records attributed to M Sekiguchi.

At least 595 records · Page 33Linked to original sources
PubMed1982

Are purine bases enzymatically inserted into depurinated DNA in Escherichia coli?

An enzyme activity to incorporate labeled dATP or dGTP preferentially into depurinated DNA, found in an extract of Escherichia coli, does not seem to represent "purine insertase," but may be ascribed to a combined action of DNA polymerase I and AP endonuclease(s) on the basis of the following findings. (1) The activity was found in polA+ but not in polA-cells. (2) The base moiety and the alpha-phosphate group of the nucleotide were equally incorporated into the DNA.

DNA Polymerase I↗
PubMed1982

Long-term follow-up of electrocardiographic findings in patients with acute myocarditis proven by endomyocardial biopsy.

Sixteen cases of acute myocarditis, proven by endomyocardial biopsy, of possible viral origin which were described in a previous paper have been followed up for up to 5 years. The cases were divided into 3 groups according to the electrocardiographic (ECG) patterns: with conduction disturbance at the early stage (Group I, 9 cases), without conduction disturbance (Group II, 4 cases); early death cases in which the follow-up could not be carried out (Group III, 3 cases). In Group I, 2 cases revealed persistent complete A-V block, 4 showed incomplete recovery of right bundle branch and/or fascicular blocks, and 3 showed complete recovery of the ECG abnormalities. In Group II, all 4 cases showed a pseudoinfarction pattern which consisted of abnormal Q waves, poor R wave progression and ST elevation. These findings disappeared during the follow-up period. All patients of Groups I and II are still alive after a follow-up period of up to 5 years.

Acute Disease↗
PubMed1982

Angiographic evidence of left main trunk spasm as a possible cause of syndrome X.

The present case is a 49-year-old female who began to feel a chest pain during exercise 2 years ago. Recently, the chest pain occurred during light exercise as well as at rest. The electrocardiogram during the attack showed remarkable ischemic ST segment depression. No abnormality was found in the coronary cineangiography. She was diagnosed as Syndrome X. However, coronary cineangiography during an attack on another day showed a severe spasm in the left main trunk. Thus, a spasm must be considered as one of the causes of "Syndrome X".

Angina Pectoris, Variant↗
PubMed1982

Cloning of mutator genes and identification of their products.

To elucidate molecular mechanisms leading to the increment of mutation frequency by mutator genes, we have cloned the dnaQ and uvrD genes of Escherichia coli and identified their gene products. By in vitro recombination hybrid plasmids capable of complementing a dnaQ mutation were constructed. The dnaQ+ plasmids consist of a full-length pBR322 DNA and a 1.5-kilobase DNA fragment derived from the E. coli chromosome. Specific labeling of plasmid-encoded proteins by the maxicell method revealed that the 1.5-kilobase insert codes for two proteins, one with a molecular weight of 25,000 and the other with a weight of 21,000. Because insertion of the gamma delta sequence into the dnaQ gene of the plasmid resulted in disappearance of the 25,000-dalton protein, it was concluded that that protein is the dnaQ gene product. The 21,000-dalton protein was identified as RNase H. The uvrD (uvrE, recL, mutU, and pdeB) gene has been cloned with phage lambda as vector. The increased sensitivity to ultraviolet light, high mutability and conditional lethality of uvrD- strains and their derivatives were all suppressed by lysogenization of the mutant cells with lambda uvrD+. In addition to the uvrD gene, lambda uvrD+ carried the corA gene that controls transport of Mg2+, Mn2+, and Co2+ through the cell membrane. By analyzing proteins produced by the transducing phages, the uvrD and corA gene products were identified as a 75,000-dalton protein and a 37,000-dalton protein, respectively.

Bacterial Proteins↗
PubMed1982

Inhibition of leukocyte migration in agar by soluble extract from a human ovarian carcinoma cell line.

A soluble membrane extract was prepared by a hypotonic extraction method from a cultured cell line, CKS, derived from serous cystadenocarcinoma of the ovary. The leukocyte migration inhibition assay in agar was used to determine the cell-mediated immune response with this extract in patients with ovarian carcinoma in comparison with patients with other benign or malignant tumors and normal healthy subjects. With 100 micrograms protein/ml of the extract, leukocytes from 15/25 patients with ovarian carcinoma showed positive response, whereas 5/30 patients with benign tumors, 2/23 patients with non-ovarian carcinomas, and 0/13 normal healthy controls did so. These findings suggested that the soluble membrane extract from the CKS cells specifically inhibited migration of leukocytes from patients with ovarian carcinoma. There was a tendency that the stage III or IV patients responded to the extract more frequently than the stage I patients irrespective of histological type of ovarian carcinoma. The migration inhibition assay with a basic encephalitogenic protein (100 micrograms protein/ml) from bovine brain was performed simultaneously with the ovarian cancer extract. One of 20 patients with ovarian carcinoma, 1/29 patients with benign tumors, 1/22 patients with non-ovarian carcinomas, and 0/13 normal healthy controls responded positively. Therefore, cross-reactivity between the basic protein and tumor-associated antigens could not be demonstrated in the present study.

Adenocarcinoma↗
PubMed1981

Molecular cloning of the uvrD gene of Escherichia coli that controls ultraviolet sensitivity and spontaneous mutation frequency.

The uvrD gene of Escherichia coli that controls UV sensitivity and spontaneous mutation frequency has been cloned with phage lambda as vector. The increased sensitivity to ultraviolet light (UV) of uvrD3, uvrE502, recL152, and pdeB41 mutants, high mutability of uvrD3 and pdeB41 mutants, and conditional lethality of strain TS41 that carried pdeB41, polA1, and supl26 mutations were all suppressed by lysogenization of the mutant cells with lambda uvrD+. These results were consistent with the idea that the uvrD, uvrE, recL, and pdeB mutations are alleles of the uvrD gene. In addition to the uvrD gene, lambda uvrD+ carried the corA gene that controls transport of Mg++, Mn++, and Co++ through the cell membrane. Hybrid plasmids carrying both uvrD and corA genes were also constructed by using pKY2289 as a cloning vehicle. Orientational isomers that carried the same 12.0 kb fragment in the opposite direction were equally efficient in complementing the UvrD- as well as CorA- defects of the transformed host cells, suggesting that the DNA insert contains all the genetic signals needed to express the two gene products. Insertion of the gamma delta sequence into recombinant plasmids was performed to generate appropriate restriction endonuclease target sites in the cloned DNA fragments.

Bacteriophage lambda↗
PubMed1981

Conditional lethality of Escherichia coli strains carrying dnaE and dnaQ mutations.

A double mutant of Escherichia coli K12 which carries a conditional lethal mutator mutation, dnaQ49 (Horiuchi et al. 1978), and a DNA polymerase III-deficient mutation, dnaE486 (Wechsler and Gross 1971), was found to be more thermolabile than was either of the dnaQ49 or dnaE486 single mutants. The double mutant is able to grow at 28 degrees C but not at 30 degrees C. Under the restrictive conditions DNA synthesis, but not protein synthesis, of the double mutant was suppressed. All the other combinations of dnaQ and dnaE mutation alleles tested so far rendered the cells thermolabile. A dnaZ mutation exerted a similar effect on the dnaQ strain. However, when non-specific temperature-sensitive growth mutations were combined with the dnaQ49 mutation, no such increase in thermosensitivity was observed. There is a possibility that the product of the dnaQ gene interacts directly with the DNA replicating enzyme complex.

Alleles↗
PubMed1981

Repair of ultraviolet radiation damage in xeroderma pigmentosum cells belonging to complementation group F.

DNA-repair characteristics of xeroderma pigmentosum belonging to complementation group F were investigated. The cells exhibited an intermediate level of repair as measured in terms of (1) disappearance of T4 endonuclease-V-susceptible sites from DNA, (2) formation of ultraviolet-induced strand breaks in DNA, and (3) ultraviolet-induced unscheduled DNA synthesis during post-irradiation incubation. The impaired ability of XP3YO to perform unscheduled DNA synthesis was restored, to half the normal level, by the concomitant treatment with T4 endonuclease V and ultraviolet-inactivated Sendai virus. It is suggested that xeroderma pigmentosum cells of group F may be defective, at least in part, in the incision step of excision repair.

Adult↗
PubMed1981

Physical association of pyrimidine dimer DNA glycosylase and apurinic/apyrimidinic DNA endonuclease essential for repair of ultraviolet-damaged DNA.

T4 endonuclease V [endodeoxyribonuclease (pyrimidine dimer), EC 3.1.25.1)], which is involved in repair of UV-damaged DNA, has been purified to apparent physical homogeneity. Incubation of UV-irradiated poly(dA).poly(dT) with the purified enzyme preparations resulted in production of alkali-labile apyrimidinic sites, followed by formation of nicks in the polymer. The activity to produce alkali-labile sites was optimal in a relatively broad pH range (pH 6.0-8.5), whereas the activity to form nicks had a narrow optimum near pH 6.5. By performing a limited reaction with T4 endonuclease V at pH 8.5, irradiated polymer was converted to an intermediate form that carried a large number of alkali-labile sites but only a few nicks. The intermediate was used as substrate for the assay of apurinic/apyrimidinic DNA endonuclease activity [endodeoxyribonuclease (apurinic or apyrimidinic, EC 2.1.25.2]. The two activities, a pyrimidine dimer DNA glycosylase and an apurinic/apyrimidinic DNA endonuclease, were copurified and found in enzyme preparations that contained only a 16,000-dalton polypeptide. An enzyme fraction from cells infected with bacteriophage T4v1, a mutant that is sensitive to UV radiation, was defective in both glycosylase and endonuclease activities. Moreover, occurrence of an amber mutation in the denV gene caused a simultaneous loss of the two activities, and suppression of the mutation rendered both activities partially active. These results strongly suggested that a DNA glycosylase specific for pyrimidine dimers and an apurinic/apyrimidinic DNA endonuclease reside in a single polypeptide chain coded by the denV gene of bacteriophage T4. Because the two activities exhibited different thermosensitivity, it was further suggested that conformation of the active sites for these activities may be different.

DNA↗
PubMed1981

Identification of the dnaQ gene product and location of the structural gene for RNase H of Escherichia coli by cloning of the genes.

By in vitro recombination we have constructed hybrid plasmids capable of complementing a conditional lethal mutator mutation, dnaQ49, in Escherichia coli K12. The dnaQ+ plasmids consist of a full-length pBR322 DNA and a 1.5-kilobase DNA fragment derived from the E. coli chromosome. Specific labeling of plasmid-encoded proteins by the maxicell method revealed that the 1.5-kilobase insert codes for two proteins, one whose molecular weight is 25,000 [the 25-kilodalton (kDal) protein] and the other whose molecular weight is 21,000 (the 21-kDal protein). Because insertion of gamma delta sequence into the dnaQ gene of the plasmid resulted in disappearance of the 25-kDal protein, it was concluded that the 25-kDal protein is the dnaQ gene product. The 21-kDal protein was identified as RNase H on the basis of the following evidence. (i) Cells harboring the dnaQ+ plasmids, with or without the gamma delta insertion in the dnaQ gene, had a 5- to 7-fold higher level of RNase H activity than cells harboring pBR322. (ii) After induction of cells that are lysogenized with dnaQ+-transducing lambda phages, RNase H activity increased considerably. A similar high level of RNase H activity was observed with transducing phages whose dnaQ function was inactivated by insertion of a transposon, Tn3, into the gene, (iii) The plasmid-encoded RNase H, labeled with [35S]methionine, was purified in a manner essentially similar to that of the chromosome-encoded enzyme. These results suggest that the dnaQ gene and the structural gene for RNase H, termed gene rnh, are closely linked and located at 5 min on the linkage map.

Bacterial Proteins↗
PubMed1981

Early clinical profiles of cases with histopathologically proven acute idiopathic myocarditis and a proposal for diagnostic criteria.

The early clinical features, primary care, treatment and short-term prognosis in 15 cases of acute myocarditis where diagnostic confirmation was made by endomyocardial biopsy or autopsy were analyzed. Characteristically, idiopathic myocarditis of possible viral etiology revealed preceding symptoms which consisted of flu-like symptoms, i.e., fever, upper respiratory infection (sore throat, cough), myalgia or arthralgia, general malaise, and gastrointestinal disorders (vomiting, anorexia, nausea, abdominal pain and soft stool). A severe cardiac or generalized disease condition may follow. Depending upon the progress of intensive medical and cardiac care, the patients' prognosis is not always poor. Diagnostic criteria based upon our own experience have been constructed as a proposal.

Acute Disease↗