Effects of denervation on the contents of cholesterol and membrane systems involved in muscle contraction in rabbit fast-twitch sarcotubular system.

Denervated fast-twitch rabbit muscles were progressively losing their fresh weight and the yield of sarcotubular protein was increasing. The activity of Ca(2+)-ATPase was affected but very slightly, the basal Mg(2+)-ATPase and the Mg(2+)-ATPase/Ca(2+)-ATPase ratio however increased together with a simultaneous depression of the membrane-bound acetylcholinesterase activity. We did not observe any differences in density properties of sarcotubular fractions between control and denervated muscle. However, a relative enrichment in SM and H fraction could be seen after denervation with small changes in the content of the Ca(2+)-pump protein, increased levels of calsequestrin and cholesterol, mostly in the heavy and the SM fraction. After denervation the binding sites for 3H-PN-200-110 did not show any changes in receptor affinity, but the number of putative Ca(2+)-channels increased twice along with a depression of 3H-ouabain binding sites. We suggest that the denervation of fast-twitch muscle leads to the hypertrophy of the junctional sarcoplasmic reticulum and the T-system. Changes in the cholesterol content, in the number of putative Ca(2+)-channels and in Na+, K(+)-ATPase can affect the muscle contraction.     

Structures of the α(1-3)-galactose-containing asparagine-linked glycans of a Lewis lung carcinoma cell subline resistant toAleuria aurantia agglutinin: elucidation by1H-NMR spectroscopy

Four bi-antennary glycan fractions of theN-acetyllactosamine-type, derived from a Lewis lung carcinoma (LL₂) cell subline resistant to theAleuria aurantia agglutinin were studied by 400 MHz¹H-NMR spectroscopy. By this method, their antennae were found to be terminated either by (2-3 or 6)-linkedN-acetylneuraminic acid or (1-3)-linked galactose residues. The primary structure of glycans of these four glycopeptide or derived oligosaccharide-alditols has been determined in full detail.Abbreviations NAc N-acetyl group - NGc N-glycolyl group - GlcNAc N-acetylglucosamine - NeuAc N-acetylneuraminic acid - NeuGc N-glycolylneuraminic acid - Man mannose - Gal galactose - Fuc fucose - Con A concanavalin A - LCA Lens culinaris agglutinin - AAA Aleuria aurantia agglutinin - WGA Wheat germ agglutinin - RCA II Ricinus communis agglutinin II - PBS phosphate buffered saline, 0.01m Na₂HPO₄/0.14m NaCl, pH 7.2 - HPLC high performance liquid chromatography - EMEM Eagle's Minimal Essential Medium - Lec^R lectin resistant - MG -methylglycoside     

Ca2+ calmodulin-dependent protein kinase activity in the ascomycetes Neurospora crassa

DEAE-cellulose column chromatography of Neurospora crassa soluble mycelial extracts leads to the resolution of three major protein kinase activity peaks designated PKI, PKII, and PKIII.PKII activity is stimulated by Ca²⁺ and Neurospora or brain calmodulin. Maximal stimulation was observed at 2 µM-free Ca²⁺ and 1 µg/ml of the modulator. The stimulatory effect of the Ca²⁺-calmodulin complex was blocked by EGTA and by some calmodulin antagonists such as phenothiazine drugs or compound 48/80.PKII phosphorylates different proteins, among which histone II-A at a low concentration and CDPKS, the synthetic peptide specific for Ca²⁺-calmodulin dependent protein kinases, are the best substrates. Some phosphorylation can be detected in the absence of any exogenous acceptor. PKII activity assayed in the presence of histone II-A or in the absence of exogenous phosphate acceptor (autophosphorylation) co-elute in a DEAE-cellulose column at 0.28 M NaCl. As result of the autophosphorylation reaction of the purified enzyme a main phosphorylated component of 70 kDa was resolved by SDS-polyacrylamide gel electrophoresis. It is possible that this component is an active part of this enzyme.     

The role of NADPH in the regulation of glucose-6-phosphate and 6-phosphogluconate dehydrogenases in rat adipose tissue

Summary Previous studies examining the regulation of the synthesis of G6PDH and 6PGDH in rat liver and adipose tissue have focused on the induction of these enzymes by different diets and some hormones. In rat liver these enzymatic activities seem to be regulated by a mechanism involving changes in the NADPH requirements. In this paper we have studied the effect of changes in the flux through different NADPH-consuming pathways on G6PDH and 6PGDH levels in adipose tissue and on the NADPH/NADP ratio. The results show that: I) an increase in the consumption of NADPH, caused by the activation of either fatty acid synthesis or detoxification systems which consume NADPH, is paralleled by an increase in the levels of these enzymes; II) when the increase in consumption of NADPH is prevented, the G6PDH and 6PGDH levels do not change.Abbreviations G6PDH Glucose-6-Phosphate Dehydrogenase - 6PGDH 6-Phosphogluconate Dehydrogenase - GR Glutathione Reductase - ME Malic Enzyme - tBHP t-Butyl Hydroperoxide - NF Nitrofurantoin - CumOOH Cumene Hydroperoxide