Amino acid environment determines expression of carbamoylphosphate synthetase and phosphoenolpyruvate carboxykinase in embryonic rat hepatocytes

1985 ◽  
Vol 21 (11) ◽  
pp. 606-611 ◽  
Author(s):  
Wouter H. Lamers ◽  
Marian Roon ◽  
Piet G. Mooren ◽  
André Graaf ◽  
Robert Charles
Keyword(s):  
Amino Acid ◽  
Phosphoenolpyruvate Carboxykinase ◽  
Rat Hepatocytes ◽  
Carbamoylphosphate Synthetase ◽  
Acid Environment

scholarly journals Molecular cloning, sequencing and expression of the cDNA of the mitochondrial form of phosphoenolpyruvate carboxykinase from human liver

1996 ◽  
Vol 315 (3) ◽  
pp. 807-814 ◽  
Author(s):  
Said MODARESSI ◽  
Bruno CHRIST ◽  
Jutta BRATKE ◽  
Stefan ZAHN ◽  
Tilman HEISE ◽  
...  
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Human Liver ◽  
Phosphoenolpyruvate Carboxykinase ◽  
Amino Acid Level ◽  
Cdna Probe ◽  
Rat Hepatocytes ◽  
Eukaryotic Expression ◽  
Amino Acid Sequences ◽  
Sequence Identity

In human liver, phosphoenolpyruvate carboxykinase (PCK; EC 4.1.1.32) is about equally distributed between cytosol and mitochondria in contrast with rat liver in which it is essentially a cytosolic enzyme. Recently, the isolation of the gene and cDNA of the human cytosolic enzyme has been reported [Ting, Burgess, Chamberlain, Keith, Falls and Meisler (1993) Genomics 16, 698–706; Stoffel, Xiang, Espinosa, Cox, Le Beau and Bell (1993) Hum. Mol. Genet. 2, 1–4]. It was the goal of this investigation to isolate the cDNA of the human mitochondrial form of hepatic PCK. A human liver cDNA library was screened with a rat cytosolic PCK cDNA probe comprising sequences from exons 2 to 9. A cDNA clone was isolated which had overall a 68% DNA sequence and a 70% deduced amino acid sequence identity with the human cytosolic PCK cDNA. Without the flanking 270 bases (=90 amino acids) each at the 5´ and 3´ end, the sequence identity was 73% on the DNA and 78% on the amino acid level. The isolated cDNA had an open reading frame of 1920 bp; it was 54 bp (equivalent to 18 amino acids) longer than that of human or rat cytosolic PCK cDNA. The isolated cDNA was cloned into the eukaryotic expression vector pcDNAI and transfected into human embryonal kidney cells HEK293; PCK activity was increased by 3-fold in the mitochondria, which normally contain 70% of total PCK activity, but not in the cytosol. The isolated cDNA was also transfected into cultured rat hepatocytes; again, PCK activity was enhanced by about 40-fold in the mitochondria, which normally possess only 10% of total PCK activity, but not in the cytosol. In the rat hepatocytes only the endogenous cytosolic PCK and not the transfected mitochondrial PCK was induced 3-fold with glucagon. Comparison of the amino acid sequences deduced from the isolated cDNA with human and rat cytosolic PCK showed that the additional 18 amino acids were located at the N-terminus of the protein and probably constitute a mitochondrial targeting signal. Northern-blot analyses revealed the human mitochondrial PCK mRNA to be 2.25 kb long, about 0.6 kb shorter than the mRNA of the cytosolic PCK. Primer extension experiments showed that the 5´-untranslated region of mitochondrial PCK mRNA was 134 nucleotides in length.

Amino Acid Environment Affinity Model Based on Graph Attention Network

2021 ◽  
Author(s):  
Xueheng Tong ◽  
Shuqi Liu ◽  
Jiawei Gu ◽  
Chunguo Wu ◽  
Yanchun Liang ◽  
...  
Keyword(s):  
Amino Acid ◽  
Attention Network ◽  
Model Based ◽  
Acid Environment ◽  
Affinity Model

scholarly journals Involvement of a local Fenton reaction in the reciprocal modulation by O2 of the glucagon-dependent activation of the phosphoenolpyruvate carboxykinase gene and the insulin-dependent activation of the glucokinase gene in rat hepatocytes

1998 ◽  
Vol 335 (2) ◽  
pp. 425-432 ◽  
Author(s):  
Thomas KIETZMANN ◽  
Torsten PORWOL ◽  
Karl ZIEROLD ◽  
Kurt JUNGERMANN ◽  
Helmut ACKER
Keyword(s):  
Hydroxyl Radical ◽  
Hydroxyl Radicals ◽  
Fenton Reaction ◽  
Phosphoenolpyruvate Carboxykinase ◽  
Gene Activation ◽  
Three Dimensional ◽  
Rat Hepatocytes ◽  
Confocal Laser ◽  
Radical Scavenger ◽  
Insulin Dependent

H2O2 mimicked the action of periportal pO2 in the modulation by O2 of the glucagon-dependent activation of the phosphoenolpyruvate carboxykinase (PCK) gene and the insulin-dependent activation of the glucokinase (GK) gene. H2O2 can be converted in the presence of Fe2+ in a Fenton reaction into hydroxyl anions and hydroxyl radicals (•OH). The hydroxyl radicals are highly reactive and might interfere locally with transcription factors. It was the aim of the present study to investigate the role of and to localize such a Fenton reaction. Hepatocytes cultured for 24 h were treated under conditions mimicking periportal or perivenous pO2 with glucagon or insulin plus the iron chelator desferrioxamine (DSF) or the hydroxyl radical scavenger dimethylthiourea (DMTU) to inhibit the Fenton reaction. PCK mRNA was induced by glucagon maximally under conditions of periportal pO2 and half-maximally under venous pO2. GK mRNA was induced by insulin with reciprocal modulation by O2. DSF and DMTU reduced the induction of PCK mRNA to about half-maximal and increased the induction of GK mRNA to maximal under both O2 tensions. Hydroxyl radical formation was maximal under arterial pO2. Perivenous pO2, DSF and DMTU each decreased the formation of •OH to about 70% of control. The Fenton reaction could be localized in a perinuclear space by confocal laser microscopy and three-dimensional reconstruction techniques. In the same compartment, iron could be detected by electron-probe X-ray microanalysis. Thus a local Fenton reaction is involved in the O2 signalling, which modulated the glucagon- and insulin-dependent PCK gene and GK gene activation.

scholarly journals Properties of phosphoenolpyruvate carboxykinase (guanosine triphosphate) synthesized in hepatoma cells in the presence of amino acid analogues

1975 ◽  
Vol 146 (3) ◽  
pp. 585-593 ◽  
Author(s):  
S E Knowles ◽  
J M Gunn ◽  
L Reshef ◽  
R W Hanson ◽  
F J Ballard
Keyword(s):  
Amino Acid ◽  
Phosphoenolpyruvate Carboxykinase ◽  
Hepatoma Cells ◽  
Dibutyryl Cyclic Amp ◽  
Enzyme Degradation ◽  
Amino Acid Analogues ◽  
Enzyme Molecules ◽  
Natural Amino Acids

1. Phosphoenolpyruvate carboxykinase (GTP) was induced by a combination of dibutyryl cyclic AMP, theophyline and dexamethasone in Reuber H35 hepatoma cells under conditions where an amino acid in the medium was replaced by an appropriate analogue. 2. With canavanine replacing arginine or with 5-fluorotryptophan or 6-fluorotryptophan replacing tryptophan the induced enzyme had a lower catalytic activity-relative to antibody reactivity. 3. These aberrant enzyme molecules were heat-labile in vitro. 4. Measurements of enzyme degradation in vivo indicated that the canavanine-containing enzyme and the 6-fluorotryptophan-containing enzyme were degraded more rapidly than the enzyme containing all natural amino acids.

Sign in / Sign up

Export Citation Format

Share Document