Density distribution of 2′,3′-cyclic nucleotide 3′-phosphodiesterase and myelin proteins in particulate material from myelin deficient (mld) mutant and control brains

Total particulate material from control and myelin deficient (mld) brains was subjected to density centrifugation on a continuous sucrose gradient. Particles from control brains distributed in a bell-shaped mode with a peak density near 0.64 M-sucrose. In mld material only a slight elevation of optical density was observed near 0.8 M-sucrose. The highest specific activities of 2′,3′-cyclic nucleotide 3′-phosphodiesterase were observed at densities of 0.63 and 0.71 M-sucrose for mld and control brains, respectively. The peak of myelin basic protein in control fractions was near 0.60 M-sucrose. In mld fractions no peak was observed. Proteolipid and Wolfgram proteins had a maximum near 0.65 and 0.73 M-sucrose in control and mld fractions, respectively. The absence of myelin basic proteins in all the fractions makes it unlikely that, in mld mice, myelin basic proteins are synthesized but not incorporated into myelin.     

On the origin of mitochondria: a reexamination of the molecular structure and kinetic properties of pyruvate dehydrogenase complex from brewer''s yeast

45Ca2+ incorporated in response to glucose was selectively mobilized from the beta-cell-rich pancreatic islets of ob/ob-mice after raising the intracellular Na+ by removal of K+ or addition of ouabain or veratridine. Also studies of insulin release indicated opposite effects of glucose and Na+ on the intracellular sequestration of calcium. The fact that glucose inhibits insulin release induced by raised intracellular Na+ indicates that this sugar can lower the cytoplasmic [Ca2+]. The concept of a dual action of glucose on the cytoplasmic [Ca2+]. The concept of a dual action of glucose on the cytoplasmic [Ca2+] might well explain previous observations of an inhibitory component in the glucose action on the 45Ca2+ efflux.     

Identification of N- and C-terminal corticotropin peptides in the Mr 80000 form of neurophysin

The 125I-labeled Mr 80000 form of neurophysin has been purified from bovine neurohypophysi. Tryptic digests of this species were analyzed, prior to or after treatment with carboxypeptidase B, by high-pressure liquid chromatography followed by isoelectric focusing and the fragments compared with those generated by a similar treatment of reference bovine 1-39 adrenocorticotropin. The ACTH peptides 22-39 and 1-8, as well as the 1-7 derivative of the latter were identified by those two independent criteria. This provides chemical evidence supporting the hypothesis [8] that high Mr neurophysin may contain the sequence of ACTH.     

Solubilization and partial purification of protein kinase systems from brain membranes that phosphorylate calspectin: A spectrin-like calmodulin-binding protein (fodrin)

In brain tissue a spectrin-like calmodulin-binding protein calspectin, or fodrin, is concentrated in a synaptosome fraction, where most of the calspectin is associated with the synaptic membranes. This endogenous calspectin was phosphorylated by protein kinase system(s) associated with the membranes. Here, we report the solubilization and partial purification of the membrane-associated calspectin kinase activity. The activity was resolved on a gel filtration column into two fractions, peaks I and II having estimated M_r of 800 000 and 88 000. The activity of peak I was dependent on the presence of both Ca²⁺ and calmodulin. Peak II revealed a basal activity in the absence of Ca²⁺ and calmodulin, which was stimulated 2-fold by addition of Ca²⁺. Calmodulin had no effect on the peak II activity.     

Myosin light chain kinase binding to plastic

Methionine-81 and/or -8 of the transmembrane sialoglycoprotein, glycophorin A, have been specifically alkylated with ¹³CH₃I to produce the sulfonium ion derivatives [S-[¹³C]methylmethionine-8]glycophorin A and [S-[¹³C]methylmethionine-8 and -81]glycophorin A. ¹³C NMR spectra of these species show that the resonances of the methyl groups of the modified glycophorins occur at 26.1 ppm downfield from Me₄Si. A spin-lattice relaxation time of 0.4 was observed for the ¹³C-enriched methyl resonances of the sulfonium ion derivatives of Met-8 and -81, which corresponds to an effective correlation time of < 2× 10^?10 s. Demethylation of the 2 glycophorin A sulfonium ion species with 2-mercaptoethanol produces native glycophorin A which now has the ε-carbon of the methionine residue(s) 45% isotopically enriched. The ε-carbon of Met-8 was found to occur at 15.7 ppm downfield from Me₄Si whereas the ε-carbon of Met-81 exhibited an unusual chemical shift of 2.0 ppm downfield from Me₄Si. The spin-lattice relaxation time of both resonances was found to be ～0.3 s.     

Active sulfate absorption in rabbit ileum: Dependence on sodium and chloride and effects of agents that alter chloride transport

Summary Unidirectional fluxes of³⁵SO₄ across and into rabbit ileal epithelium were measured under short-circuit conditions, mostly at a medium SO₄ concentration of 2.4mm. Unidirectional mucosa (m)-to-serosa (s) ands-to-m fluxes (J _ms,J _sm) were 0.456 and 0.067 moles hr^–1 cm^–2, respectively.J _ms was 2.7 times higher in distal ileum than in mid-jejunum. Ouabain abolished net SO₄ transport (J _net) by reducingJ _ms. Epinephrine, a stimulus of Cl absorption, had no effect on SO₄ fluxes. Theophylline, a stimulus of Cl secretion, reducedJ _ms without affectingJ _sm, causing a 33% reduction inJ _net. Other secretory stimuli (8-Br-cAMP, heat-stable enterotoxin, Ca-ionophore A23187) had similar effects. Replacement of all Cl with gluconate markedly reducedJ _net through both a decrease inJ _ms and an increase inJ _sm. The anion-exchange inhibitor, 4-acetoamido-4-isothiocyano-2,2-sulfonic acid stilbene (SITS), when added to the serosal side, reducedJ _ms by 94%, nearly abolishingJ _net. SITS also decreasedJ _sm by 75%. Mucosal SITS (50 m) was ineffective. 4,4-diisothiocyano-2,2-sulfonic acid stilbene (DIDS) had effects similar to SITS but was less potent. Measurements of initial rates of epithelial uptake from the luminal side (J _me) revealed the following: (1)J _me is a saturable function of medium concentration with aV _max of 0.94 moles hr^–1 cm^–2 and aK _1/2 of 1.3mm; (2) replacing all Na with choline abolishedJ _me; (3) replacing all Cl with gluconate increasedJ _me by 40%; (4) serosal SITS had no effect onJ _me; and (5) stimuli of Cl secretion had no effect onJ _me or increased it slightly. Determination of cell SO₄ with³⁵SO₄ indicated that, at steady-state, the average mucosal concentration is 1.1 mmoles per liter cell water, less than half the medium concentration. Cell SO₄ was increased to 3.0mm by adding SITS to the serosal side. Despite net transport rates greater than 1.4 Eq hr^–1 cm^–2, neither addition of SO₄ to the SO₄-free medium nor addition of SITS to SO₄-containing medium altered short-circuit current. The results suggest that (1) ileal SO₄ absorption consists of Na-coupled influx (symport) across the brush border and Cl-coupled efflux (antiport) across the basolateral membrane; (2) the overall process is electrically neutral; (3) the medium-to-cell Cl concentration difference may provide part of the driving force for net SO₄ absorption; and (4) since agents affecting Cl fluxes (both absorptive and secretory) have little effect on SO₄ fluxes, the mechanisms for their transcellular transports are under separate regulation.     

The antimutagenic effect of selenium on 7,12-dimethylbenz(a)anthracene and metabolites in the amesSalmonella/microsome system

The antimutagenic effect of selenium as sodium selenite, sodium selenate, selenium dioxide, and seleno-methionine was studied in the AmesSalmonella/microsome mutagenicity test using 7,12-dimethylbenz(a)anthracene (DMBA) and some of its metabolites. Selenium (20 ppm) as sodium selenite reduced the number of histidine revertants on plates containing up to 100 μg DMBA/plate. Increasing concentrations of selenium as sodium selenite, sodium selenate, and selenium dioxide up to 40 ppm Se progressively decreased the number of revertants caused by 50 μg DMBA. DMBA and its metabolites 7-hydroxymethyl-12-methylbenz(a)anthracene, 12-hydroxymethyl-7-methylbenz(a)anthracene, and 3-hydroxy-7,12-dimethylbenz(a)anthracene were mutagenic forSalmonella typhimurium TA100 in the presence of an S-9 mixture. Selenium supplementation as Na₂SeO₃ reduced the number of revertants induced by these metabolites to background levels. The antimutagenic effect of inorganic selenium compounds cannot be explained by toxicity of selenium as determined by viability tests withSalmonella typhimurium TA100. Selenium supplementation in all forms examined, except sodium selenate, decreased the rate of spontaneous reversion. Selenium as sodium selenate was slightly mutagenic at concentrations of 4 ppm or less. Higher concentration of Na₂SeO₄ inhibited the mutagenicity of DMBA. The present studies support the anticarcinogenic potential of selenium and indicate that form and concentration are important factors in this trace element's efficacy.