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Abstract— An enzyme radiochemical assay for p -octopamine, m -octopamine (norphenylephrine) and phenylethanolamine based on the N -methylation of these amines by the enzyme phenylethanolamine N -methyl transferase ( S -adenosyl- l -methionine: phenylethanolamine N -methyl transferase (EC 2.1.1.28) has been developed. [3 H]Methyl- S -adenosyl- l -methionine was used as methyl donor. The reaction products are converted to their dansyl derivatives and separated by TLC in three different solvent systems prior to liquid scintillation counting. The method exhibits a sensitivity of less than 10 pg for each amine and is suitable for the measurement of endogenous p -octopamine levels in mammalian brain. The highest levels of p -octopamine were found in the hypothalamus (3.4 ng/g) but despite the sensitivity of the assay, neither phenylethanolamine nor m -octopamine could be detected. After MAO inhibition, however, both of these amines were found to be present. p -Octopamine was increased substantially in all brain regions following the administration of an MAO inhibitor, whereas pretreatment with reserpine produced a significant decrease in the hypothalamus. 相似文献
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The myc family of proto-oncogenes consists of several members that possess regions of sequence homology and some have known similarities in structure and function. We have isolated an 8.8-kb EcoRI fragment from a human genomic library by hybridization to a 28-base oligonucleotide probe derived from a region of the second exon of MYC, which is highly conserved in the myc gene family. Sequence analysis of this myc-like (MYCLK1) DNA fragment has revealed the existence of a region with 85% homology to the 28-base oligonucleotide probe. An open reading frame of 207 nucleotides containing the region of homology was found. We have mapped MYCLK1 to human chromosome 7 at band p15 by chromosome in situ hybridization; this site is distinct from the map location of previously characterized myc genes. Whether MYCLK1 represents a new functional member of the myc family of proto-oncogenes remains to be determined. 相似文献
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Insulin-stimulated hexose transport and glucose oxidation in rat adipocytes is inhibited by sphingosine at a step after insulin binding 总被引:1,自引:0,他引:1
D G Robertson M DiGirolamo A H Merrill J D Lambeth 《The Journal of biological chemistry》1989,264(12):6773-6779
Spingosine, a naturally occurring inhibitor of protein kinase C, has recently been shown to have potent bioregulatory effects on a variety of cellular processes involving signal transduction mechanisms. In the present studies, we have investigated its effects on activation by insulin of hexose transport and glucose oxidation in isolated rat adipocytes. Preincubation of cells with this long-chain base blocked both the marked activation of these processes by insulin and the smaller activation by phorbol myristate acetate. Inhibition of both insulin and phorbol 12-myristate 13-acetate activation showed the same sphingosine concentration dependence, suggesting a common locus of action. The effectiveness of sphingosine was inversely proportional to the lipid content in the incubation (which was a function of both the age of the animal and the number of cells used) presumably due to dilution of the lipophilic long-chain base into the cellular triglycerides. Sphingosine did not affect either insulin binding to its receptor or the half-maximal concentration of the hormone required to activate hexose transport, but reduced the maximal responses. Thus, the inhibition was at a step distal to the binding of insulin to its receptor. Basal transport activity was not inhibited, suggesting a locus of action prior to the glucose transporter. The inhibitor was also effective when added following activation by insulin of hexose transport and resulted in a rapid reversal of activation (t 1/2 for inhibition was 2-4 min.). Sphingosine and its analogs showed a parallel potency for inhibition both of isolated protein kinase C and of insulin activation in adipocytes, consistent with an essential role for protein kinase C in the activation of hexose transport by insulin. 相似文献
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