Fluorometric assay of enzymatic reactions involving acetyl Coenzyme A in aldol condensations.
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Biomedical subjects
Publications and source records attributed to J M Calvo.
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About 600 compounds were screened as possible carbon or nitrogen sources for Salmonella typhimurium LT-2. About 100 utilizable compounds were found.
5',5',5'-Trifluoro-dl-leucine inhibited the activity of alpha-isopropylmalate synthetase (the initial enzyme unique to leucine biosynthesis) as well as the growth of Salmonella typhimurium. Mutants of S. typhimurium resistant to the analogue were isolated and characterized. In most cases, they overproduced and excreted leucine or leucine, valine, and isoleucine as a result of an alteration in the regulation of branched-chain amino acid biosynthesis. Biochemical and genetic tests allowed the mutants to be grouped into three classes: I, a moderately large group (13%) which had high, constitutive leucine biosynthetic enzyme levels and mutant sites linked to the leucine operon (operator constitutive); II, a single mutant in which the mutant site was linked to the leucine operon and in which alpha-isopropylmalate synthetase was not inhibited by leucine (feedback negative); III, a majority type which had constitutive levels of leucine, valine, and isoleucine biosynthetic enzymes and mutant sites unlinked to the leucine operon. Mutants of class I provide important evidence for the concept of an operon organization of genes involved in leucine biosynthesis. The properties of class III mutants indicate that there is some element involved in regulation which is common to the three pathways.
Mutants of Salmonella have been isolated which luck either end product inhibition or repression of leucine biosynthesis. In a minimal saltsglucose medium, growth is not impaired by the lack of either control mechanism alone. The loss of both control mechanisms, however, leads to a 43 percent reduction in cell yield and a 1.5-fold reduction in the growth rate.
The effect of repetitive administration of naloxone on the development of massed amygdaloid kindling in 'encéphale isolé' cats was studied. Electrical amygdaloid kindling was carried out with a 15 min inter-stimulus interval (ISI) in a control situation with intravenous (i.v.) naloxone administration (2, 4, and 8 mg/kg), 5 min prior to amygdaloid stimulation. It was found that it was possible to complete the amygdaloid kindling process in the encéphale isolé preparation reaching generalized electrographic tonic-clonic self-sustained seizures. The enhancement of the duration, frequency, and propagation of the after-discharge (AD) was accentuated by naloxone which also induced a progressive amplitude increment of the first potential evoked by the onset of the tetanus. The number of trials needed to achieve seizure generalization was reduced in dose-dependent manner by naloxone. The ability of naloxone to accelerate the development of amygdaloid kindling may be related to an inhibitory role of opioid peptides in this process.
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In Escherichia coli K12 and in Salmonella typhimurium the first step common to the biosynthesis of isoleucine, leucine and valine is catalyzed by an intriguing system of isoenzymes. Two of these are normally expressed, while the genetic determinant for a third one is transcribed, but not translated as an active polypeptide. We analyze here the significance of this system in the light of the most recent results.