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Biomedical subjects

S Numa

Publications and source records attributed to S Numa.

At least 127 records · Page 7Linked to original sources
PubMed1978

Construction of bacterial plasmids that contain the nucleotide sequence for bovine corticotropin-beta-lipotropin precursor.

mRNA that encodes the common peptide precursor for the hormones corticotropin and beta-lipotropin was purified from the neurointermediate lobe of bovine pituitaries, and double-stranded cDNA species synthesized from this template were cloned in Escherichia coli X1776 by inserting them into the Pst I endonuclease cleavage site of the pBR322 plasmid using poly(dG)poly(dC) homopolymeric extensions. Certain of the cloned cDNA inserts contain nucleotides corresponding to the complete amino acid sequence of bovine corticotropin and a coding sequence that corresponds to at least the first portion of bovine beta-lipotropin. The nucleotide sequences coding for corticotropin and beta-lipotropin are separated on the cDNA by a 6-base-pair sequence encoding lysine and arginine, indicating that the carboxyl terminus of corticotropin is connected on the precursor peptide with the amino terminus of beta-lipotropin by these two amino acids. In addition, the cloned cDNA insert is characterized by an unusually high C+G nucleotide base content as well as by a number of DNA sequence duplications.

Adrenocorticotropic Hormone↗
PubMed1978

Effects of steroid hormones on the level of corticotropin messenger RNA activity in cultured mouse-pituitary-tumor cells.

Studies have been made with the mouse pituitary tumor cell line AtT-20 in culture to determine whether or not the suppression of pituitary corticotropin messenger RNA activity observed upon the administration of glucocorticoids to adrenalectomized rats is due to a direct action of these steroid hormones on the pituitary. The levels of corticotropin messenger RNA activity in AtT-20 cells treated with various steroid hormones were measured with the use of the cell-free protein-synthesizing system derived from wheat germ. The addition of dexamethasone to culture medium reduced the level of corticotropin messenger RNA activity to 30-40% of that in untreated cells. Corticosterone and cortisol exhibited a suppressive effect to a lesser extent. In contrast, nonglucocorticoids such as testosterone and 17beta-estradiol were essentially ineffective. These results indicate that at least part of the glucocorticoid action is exerted directly on the pituitary to suppress corticotropin messenger RNA activity.

Adrenocorticotropic Hormone↗
PubMed1977

Diacylglycerol acyltransferase from rat liver microsomes. Separation and acyl-donor specificity.

1. Diacylglycerol acyltransferase was resolved from rat liver microsomes and separated from glycerolphosphate acyltransferase and 1-acylglycerolphosphate acyltransferase. The separation was achieved by sucrose-density-gradient centrifugation of the enzyme preparation which was obtained by molecular-sieve chromatography of microsomes solubilized with a nonionic detergent, Triton X-100. 2. Although diacylglycerol acyltransferase and 1-acylglycerolphosphorylcholine acyltransferase were not separated from each other, the two acyltransferases were distinguishable with respect to heat stability and sensitivity to sulfhydryl-binding reagents. 3. Studies with the diacylglycerol acyltransferase preparation obtained have shown that this enzyme possesses a broad acyl-donor specificity, utilizing saturated, monoenoic, dienoic and tetraenoic fatty acyl-CoA thioesters efficiently. 4. A simplified assay method for diacylglycerol acyltransferase is described.

1-Acylglycerophosphocholine O-Acyltransferase↗
PubMed1977

Candida lipolytica mutants defective in an acyl-coenzyme A synthetase: isolation and fatty acid metabolism.

Mutant strains of Candida lipolytica defective in an acyl-CoA synthetase [acid:CoA ligase (AMP-forming); EC 6.2.1.3]were isolated. The mutant strains apparently exhibited no acyl-CoA synthetase activity in vitro and were, in contrast to the wild-type strain, incapable of growing in the presence of exogenous fatty acid when cellular synthesis de novo of fatty acid was blocked. However, the mutant strains grew on either fatty acid or n-alkane as a sole carbon source at rates comparable to that observed for the wild-type strain. Analysis of the fatty acid composition of the lipids from the mutant cells grown on odd-chain-length fatty acid as well as [14C]oleic acid incorporation studies have shown that the mutant cells, unlike the wild-type cells, cannot incorporate exogenous fatty acid as a whole into cellular lipids, but utilize the fatty acid that is synthesized de novo from acetyl-CoA produced by degradation of exogenous fatty acid. This finding indicates the presence of at least two acyl-CoA synthetases that activate long-chain fatty acid. One, designated acyl-CoA synthetase I, which is absent in the mutant strains, is responsible for the production of acyl-CoA to be utilized for the synthesis of cellular lipids. The other acyl-CoA synthetase provides actyl-CoA that is exclusively degraded via beta-oxidation to yield acetyl-CoA.

Candida↗
PubMed1977

Glucocorticoid effect on the level of corticotropin messenger RNA activity in rat pituitary.

In an attempt to understand the molecular mechanism underlying the depressive effect of glucocorticoids on corticotropin production, the level of corticotropin messenger RNA activity in rat pituitaries was measured with the use of the cell-free protein-synthesizing system derived from wheat germ. The large translation product of corticotropin messenger RNA was identified and quantitated by indirect immunoprecipitation with antibody against corticotropin. The level of corticotropin messenger RNA activity was increased 3- to 6-fold by adrenalectomy. Dexamethasone administration to adrenalectomized rats resulted in a marked suppression of corticotropin messenger RNA activity. Cortisol and corticosterone also exhibited a suppressive effect but were less effective than dexamethasone. In contrast, nonglucocorticoids such as progesterone and aldosterone had no suppressive effect. These results indicate that at least part of the glucocorticoid effect on corticotropin production in the pituitary is exerted at the pretranslational level.

Adrenal Cortex Hormones↗
PubMed1977

Characterization of a differentiation-stimulating factor for mouse myeloid leukemia cells.

Conditioned medium containing a differentiation-stimulating factor (D-factor) for M1 cells (myeloid leukemia cell line) was recovered over a period of several weeks by culturing mouse secondary embryo cells in serum-free medium but supplemented with bovine serum albumin. Incubation of M1 cells with the conditioned medium for 48 hr or less resulted in differentiation of 60 to 80% of the cells. From the conditioned medium, D-factor was partially purified by preparative electrophoresis on a Pevikon block and the specific activity was increased 50 approximately 60-fold. The partially purified D-factor was capable of differentiating 30% of the M1 cells at a protein concentration of 2 approximately 3 microgram/ml. Half-logarithmic plots of phagocytosis-inducing activity vs. protein concentration yielded a sigmoid dose-response curve. D-factor is considered to be a glycoprotein with a molecular weight of 40,000 approximately 50,000 and an electrophoretic mobility of alpha-globulin. It is resistant to different protein-denaturing treatments. Since the nature of D-factor differs from that of the colony-stimulating factor for mouse bone marrow cells, it appears that these two activities are due to different substances.

Animals↗
PubMed1976

Evidence that acyl coenzyme A synthetase activity is required for repression of yeast acetyl coenzyme A carboxylase by exogenous fatty acids.

The cellular content of acetyl-CoA carboxylase [acetyl-CoA:carbon-dioxide ligase (ADP-forming), EC 6.4.1.2] in Saccharomyces cerevisiae is reduced by the addition of long-chain fatty acids to the culture medium. Mutant strains of S. cerevisiae defective in acyl-CoA synthetase [acid:CoA ligase (AMP-forming), EC 6.2.1.3] were isolated and used to determine whether fatty acid itself or a metabolite of fatty acid is more directly responsible for the repression of acetyl-CoA carboxylase. Cells of the mutant strains were capable of incorporating fatty acid to an extent comparable to that observed with the wild-type strain, but they accumulated markedly more of the incorporated fatty acid in the nonesterified form than did the wild-type cells. The level of acetyl-CoA carboxylase activity in the mutants, in contrast to that in the wild-type strain, was hardly affected by the addition of fatty acids to the medium. These results indicate that the activation of exogenous fatty acid is required for the repression of acetyl-CoA carboxylase, supporting the view that the repressive effect is mediated by some compound metabolically derived from fatty acid.

Acetate-CoA Ligase↗
PubMed1976

Dietary and hormonal regulation of the content of acetyl coenzyme A carboxylase-synthesizing polysomes in rat liver.

Specific polysomes involved the the synthesis of acetyl-CoA carboxylase [acetyl-CoA: carbon-dioxide ligase (ADP-forming), EC 6.4.1.2] have been identified by the binding of 125I-labeled antiacetyl-CoA carboxylase to rat liver polysomes. The binding is highly specific and occurs through the recognition of the nascent peptide chains on polysomes. With the use of the 125I-labeled antibody binding technique, it has been demonstrated that the relative content of acetyl-CoA carboxylase-synthesizing polysomes in the liver correlates well with the rate of hepatic synthesis of the enzyme in rats subjected to different dietary conditions as well as in alloxan-diabetic rats with or without insulin treatment.

Acetyl-CoA Carboxylase↗
PubMed1976

Dietary control of triglyceride and phospholipid synthesis in rat liver slices.

1. The effect of dietary manipulation on the synthesis of triglycerides and phospholipids was investigated by determining the incorporation of labeled long-chain fatty acid or glycerol into these lipids in liver slices derived from normally fed, fasted, and fat-free refed rats. 2. Triglyceride synthesis was affected markedly by the dietary regime of the animal; the lowest rates were measured with fasted rats, and the highest ones with fat-free refed rats. 3. In contrast to triglyceride synthesis, phospholipid synthesis occured at virtually constant rates regardless of the dietary conditions. 4. Addition of large amounts of fatty acid to the incubation mixture resulted in a marked stimulation of triglyceride synthesis, whereas phospholipid synthesis was affected to a much smaller extent. 5. These results indicate that the synthesis of triglycerides and that of phospholipids are controlled independently, and that the availability of fatty acid in the cell contributes to the control of triglyceride synthesis.

Animals↗
PubMed1976

Acetyl-coenzyme-A carboxylase of Candida Lipolytica. 1. Purification and properties of the enzyme.

Acetyl-coenzyme-A carboxylase has been isolated in homogeneous form from Candida lipolytica. The homogeneity of the enzyme preparation is evidenced by analytical ultracentrifugation, dodecyl-sulfate-polyacrylamide gel electrophoresis and Ouchterlony double-diffusion analysis. The purified enzyme exhibits a specific activity of 8.0 U/mg protein at 25 degrees C and contains 1 mol biotin/263000 g protein. The sedimentation coefficient (S20,W) of the enzyme is 18 S. It has been shown by dodecyl-sulfate-polyacrylamide gel electrophoresis that the enzyme possesses only one kind of subunit with a molecular weight of 230000. This finding, together with the biotin content, indicates that the C. lipolytica enzyme has a highly integrated subunit structure. The C. lipolytica enzyme is very labile, but is stabilized by glycerol. The enzyme is markedly activated by poly(ethyleneglycol), the activation being due principally to a decrease in the Km values for substrates. Even in the presence of this activator, the Km value for acetyl-CoA of the C. lipolytica enzyme is much higher than that of the enzyme from Saccharomyces cerevisiae and animal tissues. The C. lipolytica enzyme, unlike the enzyme from animal tissues, is not activated by citrate.

Acetyl-CoA Carboxylase↗
PubMed1976

Acetyl-coenzyme-A carboxylase of Candida lipolytica. 2. Regulation of cellular content and synthesis of the enzyme.

The level of acetyl-coenzyme-A carboxylase activity in Candida lipolytica undergoes large variations depending upon the carbon source on which the yeast is grown. Cells grown on n-alkanes or fatty acids exhibit a lower activity level than do cells grown on glucose. Among the n-alkanes and fatty acids tested, n-heptadecane, n-octadecane, oleic acid and linoleic acid reduce the enzyme activity to the lowest levels, which are 16-18% of the activity level in glucose-grown cells. Immunochemical titrations and Ouchterlony double-diffusion analysis with specific antibody as well as kinetic studies have indicated that the observed decrease in the level of acetyl-CoA carboxylase activity is due to a reduction in the cellular content of the enzyme. Furthermore, isotopic leucine incorporation studies with the use of the immunoprecipitation technique have demonstrated that the relative rate of synthesis of the enzyme in oleic-acid-grown cells is diminished to 12% of that in glucose-grown cells. Evidence has also been obtained to support the view that the enzyme in this yeast is not degraded at a rate high enough to contribute to the marked decrease in the cellular content of the enzyme. Thus, it is concluded that the reduction in acetyl-CoA carboxylase content in fatty-acid-grown cells is due to diminished synthesis of the enzyme.

Acetyl-CoA Carboxylase↗