Metabolic control and compartmentation in single living cells

Microspectrofluorometry of cell coenzymes (NAD(P)H, flavins) in conjunction with sequential microinjections into the same cell of metabolites and modifiers, reveals aspects of the regulatory mechanisms of transient redox changes of mitochondrial and extramitochondrial nicotinamide adenine dinucleotides. The injection of ADP in the course of an NAD(P)H transient produced by glycolytic (e.g. glucose 6-phosphate, G6P) or mitochondrial (e.g. malate) substrate leads to sharp reoxidation (state III, Chance and Williams, 1955), followed by a spontaneous state III to IV transition, and an ultimate return to original redox steady state. The response to ADP alone is biphasic, i.e. a small oxidation-reduction transient followed by a larger reverse transient. Similarities between responses to injected ATP and ADP suggest possible intracellular interconversions. Sequential injections of glycolytic and Krebs cycle substrates into the same cell, produce a two-step NAD(P) response, possibly revealing the intracellular compartmentation of this coenzyme. A two-step NAD(P)H response to sequentially injected fructose 1,6-diphosphate and G6P indicates the dynamic or even structural compartmentation of glycolytic phosphate esters in separate intracellular pools. The intracellular regulation and compartmentation of bioenergetic pathways and cell-to-cell metabolic inhomogeneities provide the basis on which the quantitative biochemistry of the intact living cell may be reconciled with these in situ findings.     

Comparison of Isocitrate Dehydrogenase Activity, Pyruvate Kinase Activity, and Polyphenol Content in Physiologically Different Pine Callus Tissue

During the growth of callus tissue of slash pine (Pinus elliottil Engelm.) several physiologically different types of tissue can be observed, often within the same culture. Different tissues were selected, based on color appearance, and used to determine isocitrate dehydrogenase and pyruvate kinase activity, and total polyphenol content. Isocitrate dehydrogenase and pyruvate kinase activity in yellow tissue was 3- to 5-fold greater than in brown tissue, whereas the polyphenol content in yellow tissue was approximately 5-fold less than in brown tissue. Dark brown callus, which also contained large amounts of polyphenols, did not have detectable enzyme activity. The differences in optimal concentrations of substrate and cofactors for the isocitrate dehydrogenase and pyruvate kinase reactions in yellow and brown tissues were very minor and therefore cannot account for the 3- to 5-fold difference in enzyme activity between these tissues. Also, the addition of brown or dark-brown tissue extract to the yellow tissue extract did not inhibit isocitrate dehydrogenase or pyruvate kinase activity in the yellow tissue extract.     

Chlorite dismutase from Ideonella dechloratans

Chlorite dismutase has been purified from the chlorate-metabolizing bacterium Ideonella dechloratans. The purified enzyme is tetrameric, with a relative molecular mass of 25,000 for the subunit, and contains about 0.6 heme/subunit as isolated. Its catalytic properties are similar, but not identical, to those found for a similar enzyme purified earlier from the bacterium GR-1. The heme group in Ideonella chlorite dismutase is readily reduced by dithionite, in contrast to the GR-1 enzyme, and redox titration gave a value of -21 mV for the midpoint potential at pH 7. The heme group has been characterized by optical and EPR spectroscopy. It is high-spin ferric at neutral pH, with spectroscopic properties similar to those found for cytochrome c peroxidase. In the alkaline pH range, a low-spin compound is formed. A 22-residue N-terminal amino acid sequence has been determined and no homologue has been found in the protein sequence databases.     

Ontogeny and ultrastructure of somatostatin and calcitonin cells in the thyroid gland of the rat

Summary Calcitonin cells are relatively numerous in the thyroid gland of the rat. In contrast, somatostatin cells are very scarce except at the time of birth and a few days thereafter, when they are conspicuously numerous. Somatostatin cells of the thyroid gland, which are ultrastructurally similar to somatostatin cells in gut and pancreas, also contain immunoreactive calcitonin. It is not clear whether somatostatin cells in the rat thyroid gland produce calcitonin or accumulate calcitonin from the environment.     

Gut-type glucagon immunoreactivity in nerves of the rat brain.

Nerve fibers reacting with antisera demonstrating gut-type glucagon were numerous in certain areas of hypothalamus and thalamus but absent from neocortex and hippocampus. They did not react with glucagon antisera specific for pancreatic type glucagon. Immunoreactive cell bodies were not observed.