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Austin L. Shug Mary J. Schmidt Gregory T. Golden Ruggero G. Fariello 《Life sciences》1982,31(25):2869-2874
A differential distribution of carnitine was found in twelve microdissected areas of the frozen canine brain. The cerebellar cortex showed the highest level of carnitine, while the lowest amounts were found in the hippocampus and pons sections of the brain. However, the inability to demonstrate changes in carnitine levels after stimulation of the major afferent pathways in the caudate nucleus, and failure to demonstrate K+ stimulated release of carnitine in rat brain slices suggest that carnitine does not have a direct neurotransmitter role in the brain. 相似文献
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The presence of repeated elements in restriction fragments used as hybridization probes for chromosomal walking poses a major obstacle to the success of this gene-cloning strategy. This report describes a simple and rapid means of identifying restriction fragments devoid of repeated sequences and therefore useful as hybridization probes for chromosomal walking. Restriction fragments derived from a genomic DNA clone are Southern blotted and hybridized to nick-translated total genomic [32P]DNA. Fragments of the genomic clone that contain high abundance sequences (i.e., repeated elements) hybridize strongly to their nick-translated counterparts, which, due to their high copy number, comprise a significant proportion of the total genomic DNA probe. Conversely, fragments containing single-copy or low-abundance sequences do not hybridize, as their nick-translated counterparts are poorly represented in the total genomic DNA probe. These latter fragments, by virtue of their low-abundance sequences, are well suited as probes for chromosomal walking. Ensuring the absence of repeated elements in restriction fragments prior to their purification and utilization as chromosomal walking probes results in marked savings of time, effort and materials. 相似文献
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The kinetics of refolding of ribonuclease A were monitored by the return of catalytic activity and inhibitor binding at -15 degrees C in 35% methanol cryosolvent at pH* 3.0 and 6.0. Catalytic activity was measured with cytidine 2',3'-cyclic monophosphate as substrate; inhibitor binding was determined with the competitive inhibitor cytidine 2'-monophosphate. Biphasic kinetics were observed at pH* 3.0 for both return of catalytic activity and inhibitor binding. At pH* 6.0 the rate of return of catalytic activity was monophasic, whereas that of inhibitor binding was biphasic. For both inhibitor binding and catalytic activity one of the observed rates was pH-dependent. Full return of catalytic activity was obtained at the completion of the refolding process. The observations are interpreted in terms of two parallel pathways of refolding for slow-refolding ribonuclease, with several native-like, partially folded intermediate states on the minor slow-refolding pathway. Of particular note is the presence of at least one such species that has inhibitor-binding capacity but not catalytic activity. This may be rationalized in terms of the known native structure. In addition, an intermediate is postulated which has the incorrect Pro-93 conformation and only partial catalytic activity (42% of the native). The slowest observed transient is attributed to the isomerization of this proline residue and return of full catalytic activity. 相似文献