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Biomedical subjects

A Peterkofsky

Publications and source records attributed to A Peterkofsky.

At least 73 records · Page 4Linked to original sources
PubMed1985

Reconstitution of regulatory properties of adenylate cyclase in Escherichia coli extracts.

The inhibition of adenylate cyclase activity of Escherichia coli by methyl alpha-glucoside has been demonstrated in intact or in permeable cells but not in cell-free extracts. In intact or permeable cells, this inhibition is demonstrable only in strains expressing the genes for proteins of the phosphoenolpyruvate:glycose phosphotransferase system (PTS); in permeable cells, the inhibition also requires potassium phosphate. Using homogeneous proteins of the PTS, we have reconstituted in cell-free extracts many of the features of the regulated form of adenylate cyclase: (i) In the absence of K2HPO4, permeable cells have lower adenylate cyclase activity than extracts; addition of homogeneous PTS proteins to the extracts brings adenylate cyclase activity close to the level observed in permeable cells. (ii) The low activity observed in permeable cells is stimulated by potassium phosphate; this stimulation is also observed in extracts supplemented with PTS proteins and phosphoenolpyruvate. (iii) In permeable cells, potassium phosphate-stimulated adenylate cyclase activity is inhibited by methyl alpha-glucoside or pyruvate; extracts behaved similarly when supplemented with PTS proteins, K2HPO4, and phosphoenolpyruvate. Thus, the regulated form of adenylate cyclase has been reconstituted in cell-free extracts by addition of homogeneous PTS proteins.

Adenylyl Cyclase Inhibitors↗
PubMed1984

Unresponsiveness of GH cells to cyclo(histidyl-proline), a metabolite of thyrotropin releasing hormone.

Cyclo(Histidyl-Proline) is a metabolite of thyrotropin-releasing hormone. It has been suggested that this peptide plays a role in regulating prolactin secretion in GH cells. An investigation of the effect of cyclo(His-Pro) on GH cells indicated that it does not affect basal prolactin release or accumulation or the levels stimulated by TRH. cAMP levels in GH cells are elevated by TRH or VIP, but not influenced by cyclo(His-Pro). cGMP levels in GH cells are not affected by either TRH or cyclo(His-Pro). While there is specific binding of TRH to receptors in GH cells, no such receptors for cyclo(His-Pro) are detectable. It is suggested that GH cells are unresponsive to cyclo(His-Pro).

Animals↗
PubMed1983

Methylglyoxal-mediated growth inhibition in an Escherichia coli cAMP receptor protein mutant.

Under certain growth conditions, some strains of Escherichia coli accumulate toxic levels of methylglyoxal. This report characterizes a strain which synthesizes a mutant cAMP receptor protein in an adenylate cyclase deletion background. When cultured in glucose 6-phosphate minimal medium, this strain (222) was prematurely growth arrested due to methylglyoxal production; growth inhibition did not occur when the strain was grown in glucose minimal medium. A comparison of a variety of enzyme and cofactor levels in the related strains 222 (mutant) and 225 (wild-type) grown on either glucose or glucose 6-phosphate medium was carried out. The only difference found that might explain an increase in methylglyoxal accumulation was an elevated level of phosphofructokinase in strain 222 grown on glucose 6-phosphate. Since this enzyme activity probably limits hexose phosphate metabolism, it is suggested that growth inhibition in strain 222 may be due to increased production of triose phosphate, some of which is converted to methylglyoxal.

Adenylyl Cyclases↗
PubMed1983

Detection in human serum by radioimmunoassay of histidyl-proline diketopiperazine, a metabolite of thyrotropin-releasing hormone.

Cyclo(His-Pro) is believed to be a metabolite of TRH. A specific antiserum directed against cyclo(His-Pro) was used to detect immunoreactive material in human serum. Gel filtration column chromatography was used to establish that cyclo(His-Pro)-like immunoreactivity was found not only in free form established to be identical to cyclo(His-Pro) by high pressure liquid chromatography analysis on reverse phase and cation exchange columns, but also in a fraction of about 100,000 mol wt. Free cyclo(His-Pro) was released by heating column fractions of 70,000 mol wt that were not immunoreactive before heating. A procedure for estimation of the content of free cyclo(His-Pro) in human serum was developed. Normal levels of cyclo(His-Pro) determined by this procedure were in the range of 11-33 pmol/ml. The level of cyclo(His-Pro) in sera of individuals with hypothyroidism, hyperthyroidism, or alcoholic cirrhosis was in the normal range, while patients with renal failure had approximately 3-fold elevated levels of the peptide. Analysis of human sera drawn at 3-h intervals over a 24-h period suggested a circadian rhythm of cyclo(His-Pro) levels.

Animals↗
PubMed1982

Histidyl-proline diketopiperazine: its biological role as a regulatory peptide.

Histidyl-proline diketopiperazine [cyclo(His-Pro)] is a metabolite of thyrotropin releasing hormone (TRH). This review summarizes the literature concerning cyclo (His-Pro) and, in addition, some studies dealing with TRH and other peptide that are considered of interest. The enzymes concerned with the metabolism of TRH are discussed. Distribution studies of peptides by immunological methods show that, while TRH is concentrated in synaptosomes, cyclo (His-Pro) is not, suggesting that cyclo (His-Pro) is not a classical neurotransmitter. Rat brain contains approximately three times as much cyclo (His-Pro) as TRH, mainly localized in the pituitary and hypothalamus. While the TRH is found in a free form, the cyclo (His-Pro) is bound to a carrier of molecular weight approximately 70,000. While specific membrane receptors for TRH have been detected in pituitary cells, no such receptors for cyclo (His-Pro) have yet been found in brain or pituitary; however, there is a specific binding of cyclo (His-Pro) to adrenal cortex membranes. Both TRH and cyclo (His-Pro) have effects in the central nervous system or pituitary. These include effects on prolactin release, thermoregulation, CNS depression, stereotypic behavior and cyclic nucleotide levels. Possible mechanisms and interrelations of these effects are discussed.

Animals↗
PubMed1981

Stimulation of Escherichia coli adenylate cyclase by lactose in strains carrying mutations in lactose permease.

When a wild-type strain of Escherichia coli contains lactose permease, the accumulation of cyclic AMP (cAMP) by intact cells is inhibited by lactose. This inhibitory effect of lactose is observed in a strain with a mutant cAMP phosphodiesterase and therefore involves a regulation of adenylate cyclase activity. Some E. coli strains carrying mutations in lactose permease show an effect opposite to that of the wild-type strain; the accumulation of cAMP by intact cells is stimulated by lactose, but only when the mutant permease is present. Insertion of lactose permease into the membrane of cells can produce a change in the specific activity of adenylate cyclase; induction of the wild-type transporter is correlated with a decrease in the specific activity, while implantation of a mutant form of lactose permease can lead to an increase in the specific activity. From these data, it is suggested that the state of the lactose transporter in the cell membrane influences the activity of adenylate cyclase.

Adenylyl Cyclases↗
PubMed1981

Escherichia coli adenylate cyclase as a sensor of sugar transport function.

Adenylate cyclase of E. coli is a membrane-bound enzyme the function of which is to synthesize a cofactor for processes that are important in metabolic transitions. The depletion from the environment of a supply of a preferred carbon source dictates the requirement for initiating the synthesis of a new metabolic system; this synthesis will require cAMP. After the adaptation period, the requirement for a high level of synthesis diminishes, resulting in a diminished requirement for cAMP. A mechanism for regulating the activity of adenylate cyclase accomplishes the variation in the required cellular cAMP concentrations. In the absence of a transportable carbon source, adenylate cyclase activity is activated by cellular regulators; when carbon sources are transported, the cellular activators are dissipated, resulting in inhibition of adenylate cyclase activity. This scheme is summarized in Fig. 6. Sugar transport systems fall into two categories: one in which the energy for the process comes from PEP (the PTS) and one in which the energy comes from the proton electrochemical gradient. Adenylate cyclase communicates with both of these systems by interacting with intermediates on the pathway to energy generation for driving these two transport processes. Adenylate cyclase couples indirectly to a large array of sugar-specific transport systems by interacting with intermediates common to all the processes. The net result of this regulatory mechanism is that, without physically communicating with the extracellular environment by spanning the membrane, adenylate cyclase effectively senses the presence of external sugars that interact with cells that have become competent to transport them.

Adenylyl Cyclases↗
PubMed1980

The subcellular and organ distribution and natural form of histidyl-proline diketopiperazine in rat brain determined by a specific radioimmunoassay.

Histidyl-proline diketopiperazine is produced in brain as a product of the metabolism of thyrotropin-releasing hormone. A number of the previously observed central nervous system and pituitary activities resulting from an exposure to thyrotropin-releasing hormone appear to involve the conversion of the releasing factor to the cyclic dipeptide. In the present study, the development of a rabbit antiserum that is highly specific for histidyl-proline diketopiperazine is described; the antiserum has essentially no capability to bind thyrotropin-releasing hormone or a number of other related peptides. The antibody can also distinguish between the natural form of the cyclic dipeptide and a diastereomer containing D-proline. A procedure for extraction, with high yield, of histidyl-proline diketopiperazine from brain is described. With the aid of the specific antiserum it was found that the preponderance of the cyclic dipeptide in rat brain is bound to high molecular weight material, mainly in the range of Mr = 70,000; histidyl-proline diketopiperazine can be disassociated from this material by boiling in salt/methanol solution. The concentration of the dipeptide in rat brain is in the range of 275 to 565 pmol/brain, approximately 2.5 times the concentrations determined for thyrotropin-releasing hormone (113 to 210 pmol/brain). A study of the subcellular distribution of histidyl-proline diketopiperazine and thyrotropin-releasing hormone suggests that the releasing factor is concentrated in synaptosomal vesicles while the diketopiperazine is not. A determination of the regional distribution of thyrotropin-releasing hormone and histidyl-proline diketopiperazine indicated that both peptides are found in highest concentrations in pituitary and hypothalamus, but are detectable in other areas of brain as well.

Animals↗
PubMed1979

Metabolism of thyrotropin releasing hormone in brain extracts. Isolation and characterization of an imidopeptidase for histidylprolineamide.

An extract of porcine brain acetone powder incubated with thyrotropin-releasing hormone (TRH; pGlu-His-ProNH2) produces acid TRH (pGlu-His-Pro), histidine, and prolineamide. Fractionation of the brain extract by DEAE-cellulose chromatography produces three protein fractions which metabolize TRH. The activity of these fractions was characterized using TRH with a 3H-label on the histidine or proline as well as [His-3H]His-ProNH2. Fraction I contains pyroglutamate aminopeptidase and Fraction II contains TRH deamidase. Fraction III was found to contain a previously unrecognized enzyme which cleaves His-ProNH2 to histidine and proline. The histidylprolineamide imidopeptidase has been characterized. A competition study using a variety of compounds containing histidine or proline suggests that the best substrates for the imidopeptidase contain a free alpha-amino group on histidine and a blocked carboxyl group on proline, as is found in His-ProNH2. A survey of a variety of polypeptide hormones indicates that many of them inhibit the imidopeptidase activity. A kinetic study of the inhibition of the enzyme by adrenocorticotropic hormone (1-24) shows that the inhibition by polypeptide hormones is noncompetitive. We hypothesize that pituitary hormones may stimulate the production of (cyclo)-His-Pro by inhibiting alternate routes of TRH metabolism.

Animals↗
PubMed1979

Escherichia coli adenylate cyclase complex: regulation by the proton electrochemical gradient.

Sugars such as glucose are transported into Escherichia coli by a coupled phosphorylation mechanism (the phosphoenolpyruvate:sugar phosphotransferase system, PTS). Transport of sugars through the PTS results in inhibition of adenylate cyclase [ATP pyrophosphate-lyase (cyclizing), EC 4.6.1.1] activity by a mechanism involving a change in the state of phosphorylation of PTS proteins. Other sugars (e.g., lactose) are transported without modification by a mechanism involving proton cotransport, which requires a proton motive force across the cell membrane. We show here that uptake of sugars through the lactose transport system results in inhibition of adenylate cyclase activity if the proton symport mechanism is also active. The protonophore carbonyl cyanide m-chlorophenylhydrazone also inhibits adenylate cyclase activity. These data suggest that the steady-state electrochemical proton gradient regulates the activity of adenylate cyclase. We propose that sugar-dependent inhibition of adenylate cyclase activity may occur by either of two mechanisms. Sugars transported by the PTS inhibited adenylate cyclase activity by dephosphorylation of a regulatory protein, while sugars transported by the proton motive force system inhibit adenylate cyclase activity as a result of collapse of the proton electrochemical gradient.

Adenylyl Cyclases↗
PubMed1978

The Escherichia coli adenylate cyclase complex: activation by phosphoenolpyruvate.

A model for the regulation of the activity of Escherichia coli adenylate cyclase is presented. It is proposed that Enzyme I of the phosphoenolpyruvate:sugar phosphotransferase system (PTS) interacts in a regulatory sense with the catalytic unit of adenylate cyclase. The phosphoenolpyruvate (PEP)-dependent phosphorylation of Enzyme I is assumed to be associated with a high activity state of adenylate cyclase. The pyruvate or sugar-dependent dephosphorylation of Enzyme I is correlated with a low activity state of adenylate cyclase. Evidence in support of the proposed model involves the observation that Enzyme I mutants have low cAMP levels and that PEP increases cellular cAMP levels and, under certain conditions, activates adenylate cyclase, Kinetic studies indicate that various ligands have opposing effects on adenylate cyclase. While PEP activates the enzyme, either glucose or pyruvate inhibit it. The unique relationships of PEP and Enzyme I to adenylate cyclase activity are discussed.

Adenosine Triphosphate↗