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Biomedical subjects

G Koike

Publications and source records attributed to G Koike.

At least 55 records · Page 3Linked to original sources
PubMed1990

Purification, structure, and biochemical properties of human O6-methylguanine-DNA methyltransferase.

The level of O6-methylguanine-DNA methyltransferase activity in a human cell line carrying a 1.1-kilobase cDNA fragment was about 50 times higher than that found in ordinary methyltransferase-proficient (Mer+) cell lines (Hayakawa, H., Koike, G., and Sekiguchi, M. (1990) J. Mol. Biol. 213, 739-747). Taking advantage of this overproduction, the enzyme was purified to apparent physical homogeneity and the physical and biochemical properties investigated. A single polypeptide with a molecular weight of approximately 25,000 was detected on sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the most highly purified preparation. The Stokes radius of 22.5 A and the sedimentation coefficient of 2.0 S were obtained, from which the molecular weight of the native form of the enzyme was calculated to be 19,000. After digestion with lysyl endopeptidase, peptide fragments of the protein were isolated and sequenced. The amino acid sequences of these peptides and the amino acid composition of the protein were in good agreement with those deduced from the nucleotide sequence of the cloned cDNA. The purified enzyme catalyzed transfer of methyl groups from O6-methylguanine and O4-methylthymine, but not from methylphosphotriesters, of methylated DNA to the enzyme molecule.

Amino Acid Sequence↗
PubMed1990

Expression and cloning of complementary DNA for a human enzyme that repairs O6-methylguanine in DNA.

A cell line with an increased resistance to alkylating agents and an extremely high level of O6-methylguanine-DNA methyltransferase activity was isolated after transfection of methyltransferase-deficient Mer- cells with a cDNA library, prepared from methyltransferase-proficient human Mer+ (Raji) cells. Sodium dodecyl sulfate/polyacrylamide gel electrophoresis analysis revealed that a protein, with a molecular weight of approximately 25,000, accepted 3H label from DNA that had been treated with [3H]methylnitrosourea. Since the cDNA for methyltransferase was integrated into the chromosomal DNA, it was recovered by using the polymerase chain reaction. When the cDNA placed in an expression vector p500 was introduced into Mer- cells, the cells acquired an increased resistance to alkylating agents and exhibited a high level of O6-methylguanine-DNA methyltransferase activity. From the transformants the cDNA could be recovered as a part of the autonomously replicating plasmid. The nucleotide sequence of the cDNA was determined, and an open reading frame comprising 207 amino acid residues was found. The molecular weight of methyltransferase, calculated from the predicted amino acid sequence, was 21,700. The predicted amino acid sequence of the human methyltransferase exhibits an intensive homology with those of the bacterial counterparts, Ada and Ogt proteins of Escherichia coli and Dat protein of Bacillus subtilis, especially around possible methyl acceptor sites.

Base Sequence↗
PubMed1990

Urinary calcium and calcium balance in young women affected by high protein diet of soy protein isolate and adding sulfur-containing amino acids and/or potassium.

The effects of sulfur-containing amino acids (SAA) and potassium (K) on urinary excretion and retention of calcium (Ca) of 27 young Japanese women were studied. A basal diet low in protein level (50 g per day) was fortified by meat or soy protein isolate (SPI) to a protein level of 100 g per day, and effects of addition of apple to these high protein diets, and addition of SAA and/or potassium (K) to the high SPI diet, especially on urinary Ca excretion, were studied. The addition of meat which increased protein intake to 100 g caused the increase in apparent absorption and urinary excretion of Ca with increased excretion of urinary sulfur (S), phosphate, ammonia, and titratable acids (TA), whereas addition of SPI did not. The addition of apple to high meat diet decreased absorption and urinary excretion of Ca. Urinary Ca, S, K, ammonia, and TA excretion increased by the addition of SAA to high SPI diet in a manner similar to the meat diet. Consequently, SAA-supplemented diet had a significantly negative effect on Ca retention. In SPI+SAA,K diet period, urinary K excretion markedly increased, and increments in urinary Ca, ammonia, and TA excretion were reversed. These changes observed in SPI+SAA, K diet period were similar to those by adding apple to meat diet without any effect on Ca absorption. The results suggest that the hypercalciuria induced by high meat diet is mainly caused by high content of SAA and may be reversed by the ingestion of K-rich foodstuffs, and soy protein does not induce hypercalciuria because of it contains less SAA than animal protein.

Absorption↗
PubMed1989

[Thrombocytosis in chronic myelogenous leukemia (CML) controlled by interferon alpha (IFN-alpha)].

A 39-year-old Japanese female who had been followed as chronic myelogenous leukemia (CML) since 1984 was admitted to our hospital because of dizziness. On admission, platelet count markedly increased (245 X 10(4)/microliters) in spite of daily administration of busulfan 2 mg. She was diagnosed as accelerated phase CML with thrombocytosis. So we tried to use interferon alpha (IFN-alpha) finally given in a dose of 9 X 10(6) U daily by subcutaneous injection. After that, platelet count decreased to 70 X 10(4)/microliters and megakaryocyte count in bone marrow decreased from 887.5/microliters to 395.7/microliters. But we had to stop IFN-alpha because of severe side effects.

Adult↗
PubMed1988

Expression of the ada gene of Escherichia coli in response to alkylating agents. Identification of transcriptional regulatory elements.

Ada protein plays a central role in the regulatory synthesis of DNA repair enzymes, following exposure of Escherichia coli to alkylating agents. Methyl groups of alkylated DNA are transferred to Ada protein by its own methyltransferase activity and the methylated Ada protein then acts as a positive regulator to overproduce the ada and related gene products. To elucidate regulatory mechanisms for the expression of the ada gene by its own product, we analyzed the ada promoter region by random and site-directed mutagenesis. A series of deletion analyses revealed that a sequence up to 53 nucleotides upstream from the transcription initiation site is required for the controlled expression of the ada gene. Libraries of base substitution mutants were constructed by synthesizing oligonucleotides corresponding to the ada promoter region in the presence of a small amount of all possible sets of nucleotides. Internal deletion and insertion mutants were also constructed with the use of synthetic oligonucleotides. Using these mutants, the -10 and the -35 boxes of the promoter as well as the ada regulatory sequence were identified, the latter being an eight-nucleotide sequence, AAAGCGCA. A six-nucleotide stretch between the regulatory sequence and the -35 box, also affected levels of expression of the gene. When the promoter DNAs derived from wild type or base substitution mutants that showed normal expression in vivo were used as templates for transcription in vitro, the ada-specific RNA was formed in the presence of a methylated form of Ada protein. With the DNAs derived from mutants of defective type as templates, no or relatively small amounts of the RNA were synthesized. Some base substitution mutants showed a constitutive expression of the gene in vivo, but this observation did not reconcile with findings in experiments in vitro.

Alkylating Agents↗
PubMed1988

Utilization and requirement of dietary protein taking into account the dermal and miscellaneous nitrogen losses in Japanese women.

Utilization and requirement of mixed protein in the conventional Japanese diet and the obligatory integumental and miscellaneous nitrogen losses were measured in female subjects. Twelve female students aged 18 to 24 years were given conventional low-protein diets at N intake levels of 50, 70, and 90 mg/kg/day for 10 days. Constituents of the diets corresponded to those of average intake of the Japanese in recent years (1982). N balances were estimated and regression equations between N intake (X) and N balance (Y) were calculated by the multiple level-constant variation method. The equation was Y = 0.426X - 40.0 (n = 36, r = 0.615, p less than 0.01) and the intersection of the regression line with zero nitrogen balance was 93.9 +/- 14.3 mg N/kg/day. The mean digestibility was 92.2 +/- 4.7%. In another experimental group, nitrogen losses due to hair, nails, and menstruation in ten Japanese women were 0.624 +/- 0.172, 0.020 +/- 0.005, and 1.76 +/- 0.68 mg/kg/day, respectively. In a third experimental group, mean of the integumental N losses was 2.8 mg/kg/day in both the subjects given a low-protein diet (19 women) and an ordinary-protein diet (4 women). It increased to 12.7 mg N/kg/day when subjects exercised in summer (4 subjects). The protein requirement in the conventional Japanese diet estimated as the sum of the mean requirement of dietary protein and obligatory dermal and miscellaneous nitrogen losses in Japanese women was 99.1 mg N/kg/day or 0.62 g protein/kg/day. The net protein utilization (NPU) of the proteins at the N intake level for N equilibrium was estimated as 48.

Adolescent↗
PubMed1986

Effect of methionine supplementation of a soy protein isolate on short-term nitrogen balance in young women.

The effect of L-methionine supplementation on the utilization of a soy protein isolate (SPI) was evaluated by short-term nitrogen balance studies in young women. Thirteen female students were given SPI in an initial period and SPI supplemented with 1% methionine in a second period immediately after menstruation as the sole source of protein. After one day on protein-free diet, each subject received conventional low-protein diet for three days, and then low protein, semisynthetic diet containing 0.5 g/kg/day (seven subjects) or 0.3 g/kg/day (six subjects) of SPI or SPI supplemented with methionine for seven days. The energy intake was approximately of a maintenance level of 36.5 +/- 3.8 kcal/kg/day. The mean N balances of the subjects at an intake level of 0.5 g/kg/day in the SPI period and methionine supplemented period were -6.2 +/- 12.6 mg N/kg and -9.8 +/- 9.8 mg N/kg, respectively, while their N balances at an intake level of 0.3 g/kg/day were -17.8 +/- 7.2 mg N/kg in the SPI period and -15.5 +/- 3.0 mg N/kg in the methionine supplemented period. There was no significant difference between the values in the SPI and methionine supplemented periods at both levels of protein intake. Blood analyses were carried out before and after the SPI period and after the period of methionine supplementation. The urinary creatinine and urea excretions during these periods were not markedly affected.

Adolescent↗
PubMed1986

Effect of fiber on protein, fat and calcium digestibilities and fecal cholesterol excretion.

Five female subjects were given four types of test diets containing various levels of protein for four consecutive 5-day periods and their dietary fiber and feces were collected throughout the experimental period. Diet A was a high-fiber, low-protein diet containing brown rice. Diet B was a semi-purified, low-protein diet containing agar agar as the sole source of dietary fiber. Diet C was a low-fiber, normal-protein diet containing polished rice. Diet D was a high-fiber, normal-protein diet containing brown rice. A fecal marker was given at breakfast on the first day of each five-day test period and on the day after the end of the experiment. Fecal weight increased during the period on high-fiber diets (diets A and D). The apparent digestibilities of protein and fat were significantly depressed by high-fiber diet. Fecal excretion of calcium did not increase on the high-fiber diets. A decrease in the apparent digestibility on a high-fiber, low-protein diet was partly due to the low intake of calcium during this period. Fecal excretion of cholesterol increased markedly during the periods on high-fiber diets. The difference between the intake and fecal excretion of dietary fiber suggested that the fiber was partially digested in the colon.

Adolescent↗
PubMed1985

Utilization and requirement of egg protein in Japanese women.

Utilization and requirement of egg protein in Japanese women with two levels of energy intake were estimated. In experiment 1, fifteen female students were given semi-purified diet containing whole egg as the sole nitrogen source for 12 days. Nitrogen intakes were 50 for five, 75 for two and 100 mg N/kg for eight subjects. Habitual energy intake was determined individually by detailed inquiry about the foods consumed before the experiment was started. Mean energy intake was 33 kcal/kg. In experiment 2, eighteen other subjects were given the same diet containing four intake levels of egg protein (30, 50, 75, 100 mg N/kg) with an energy intake of about 100 kcal/day added to that calculated by the food intake inquiry. The mean energy intake was 37 kcal/kg. The total nitrogen contents of the experimental diet, urine and feces were analyzed and the nitrogen balance was estimated from these figures. Significant rectilinear relations were found between nitrogen intake (X, mg N/kg) and balance (Y, mg N/kg). The regression equations were: Experiment 1: Y = 0.256X - 34.4 (n = 15, r = 0.742) Experiment 2: Y = 0.326X - 29.7 (n = 18, r = 0.645) The maintenance intakes of whole egg protein for apparent nitrogen equilibrium were calculated to be 134 and 91 mg N/kg with energy intakes of 33 and 37 kcal/kg, respectively. Net protein utilization (NPU) and digestibility of egg protein were calculated using the obligatory N losses previously determined in Japanese women. The NPUs at the maintenance level of egg protein with energy intakes of 33 and 37 kcal/kg were estimated as 31 and 47, respectively. The mean digestibility of egg protein was 96%.

Adolescent↗
PubMed1985

Utilization of soy protein isolate mixed with rice protein in Japanese women.

Utilization and requirement of soy protein isolate (SPI) and SPI-rice combination were examined in twenty-five female students. After 1 day on protein-free diet, each subject received a low-protein diet for 10 days. The protein sources were SPI for ten subjects and SPI-rice combination, in which the ratio of two proteins was 6:4, for fifteen subjects. The nitrogen intakes were about 45, 65 and 85 mg/kg in both the two series of experiments. Energy intake was at an approximate maintenance level of 36.1 +/- 3.0 kcal/kg. Apparent nitrogen balance improved with increase in nitrogen intake. The regression equations between nitrogen intake (X, mg/kg) and balance (Y, mg/kg) are shown in the following: SPI: Y = 0.411 X - 40.8 (n = 10, r = 0.812) SPI and rice protein: Y = 0.392 X -32.7 (n = 15, r = 0.739) From the above equations, the maintenance intakes of SPI and SPI-rice combination for an apparent nitrogen equilibrium were calculated to be 99 and 83 mg N/kg, respectively. Digestibilities were 98.2 +/- 5.0% for SPI and 93.1 +/- 6.1% for SPI-rice combination. The NPUs of SPI at intake levels of 40, 60 and 80 mg N/kg were 47 +/- 24 (n = 4), 49 (n = 2) and 44 +/- 3 (n = 4), respectively. The NPUs of SPI and rice mixed protein at intake levels of 45, 70 and 90 mg N/kg were 67 +/- 13 (n = 5), 51 +/- 7 (n = 5) and 54 +/- 12 (n = 5), respectively. It was concluded from the present study that both SPI and the SPI-rice combination had a high nutritive efficiency comparable with that of egg protein.

Adolescent↗
PubMed1984

Medical and nutritional surveys in the Kingdom of Tonga; comparison of physiological and nutritional status of adult Tongans in urbanized (Kolofo-ou) and rural (Uiha) areas.

The physiological and nutritional status of adult Tongans in rural (Uiha, 50 males and 58 females) and urbanized (Kolofo-ou, 77 males and 71 females) areas were surveyed in 1977 and 1979, respectively. Adult Tongans of both sexes in the two districts had considerably large physiques. Being different from the obesity found in peoples of industrialized countries, the obese state of adult Tongans was associated with large muscularity, low incidence of glucosuria, ECG abnormalities and hypertension, and normal plasma cholesterol levels. However, modernization has started to have an influence upon the health of adult Tongans; relatively high levels in parameters relating to obesity as well as higher incidence of hypertension were observed in adult Tongans in Kolofo-ou as compared to adult Uiha islanders.

Adult↗
PubMed1984

Effect of protein intake level on urinary energy/nitrogen ratio in Japanese.

Urinary energy/nitrogen ratios were determined in 179 female and 14 male subjects given protein from various sources and at various intake levels. The ratio decreased with increasing protein intake from zero to 1 g/kg/day but was constant when protein intake was between 1 to 1.8 g/kg/day. The ratio was not affected by the variety of protein source. There was no difference between the data for semisynthetic diet and conventional diet. Mean values and standard deviations of the ratio in men and women given the diet containing 1.0 to 1.8 g protein/kg/day were 9.06 +/- 0.56 (n = 14) and 8.19 +/- 0.81 (n = 37) kcal/kg N, respectively. The difference between two figures in men and women was significant (p less than 0.05). The mean values of urinary E/N ratio actually measured did not approach those of urea (5.34 kcal/g N), the principal nitrogenous compound in urine, the proportion of which increased at higher protein intake level. Characteristically high ratios were obtained in the ma-konbu (Laminaria japonica) and enokitake (Flammulina velutipes) diet groups. The results suggest that urinary energy originates not only from nitrogen-containing compounds but also from other organic compounds containing no nitrogen. Therefore, further investigation is necessary to evaluate the urinary E/N ratio applicable to the urinary loss of incompletely oxidized nitrogenous compounds.

Dietary Proteins↗
PubMed1983

Obligatory N loss and utilization of egg and rice mixed protein in young japanese women.

Obligatory urinary and fecal N losses in young Japanese women were evaluated and the effect of energy intake on the utilization of rice and egg mixed protein was investigated in subjects receiving a low protein diet. Seven female students were given a protein-free diet for ten days. Feces and 24-h urine samples were collected throughout the period and their nitrogen contents were analyzed. Basal metabolism was measured and the lean body mass was calculated from urinary creatinine excretion by the equation of Forbes. The mean weight loss of the 7 subjects during the ten days was 1.5 kg, the mean obligatory urinary N loss was 32.3 +/- 5.9 mg/kg body weight, 1.67 +/- 0.45 mg/kcal basal metabolism, or 1.68 +/- 0.32 g/g creatinine, and the mean obligatory fecal N loss was 10.1 +/- 2.0 mg/kg body weight, or 0.51 +/- 0.08 mg/kcal basal metabolism. In a second experiment, 17 female subjects were divided into three groups with an energy intake of about 35, 42, 46 kcal/kg BW, respectively. During the 7-day experimental period, they were given a low protein diet containing 250 g of rice and 125 g of whole egg. The nitrogen contents of the 24-h urine samples and feces collected were analyzed and the nitrogen balance was calculated as the difference between N intake and urinary and fecal N excretion. Subjects with an energy intake of 35 kcal/kg BW had a negative N balance, while subjects with an intake of 46 kcal/kg BW achieved an apparent positive N balance. The NPU values of rice and egg mixed protein with energy intake of 35, 42, 46 kcal/kg BW were calculated from the N balance data and the values for obligatory urinary and fecal N losses in the first experiment as 25, 37 and 54, respectively.

Adult↗