Membrane binding characteristics of choline-o-acetyltransferase in rat hippocampal nerve terminals
| dc.creator | Smith, L. Keith | |
| dc.date.available | 2011-02-18T20:45:20Z | |
| dc.date.issued | 1992-12 | |
| dc.degree.department | TTUHSC -- Pharmacology and Neuroscience | en_US |
| dc.description.abstract | Choline-0-acetyltransferase (ChAT; EC 2.3.1.6.) is the enzyme which catalyzes the formation of the excitory neurotransmitter acetylcholine. Although most of the ChAT in central cholinergic nerve terminals is soluble, some is also non-ionically associated with membranes. The major objective of this dissertation research was to determine how this membrane-bound detergent soluble form of ChAT (D-ChAT) was anchored to membranes in rat hippocampal nerve terminals. I tested the hypothesis that D-ChAT was anchored to membranes by a glycosylphosphatidylinositol (GPI) anchor. ChAT appeared to possess three characteristics common to many GPI-anchored proteins: (1) phosphatidylinositol specific-phospholipase C (PI-PLC) selectively released it from membranes, (2) PI-PLC treatment converted it from a detergent soluble into an aqueous form, and (3) an antibody to an epitope on the GPI anchor (anti-CRD) immunoreacted with the cytosolic form of ChAT (S3-ChAT) on Western blots. I also tested the possibility that D-ChAT might be one of the first intracellular GPI-anchored proteins described by internalizing PI-PLC into synaptosomes and determining if it would release D-ChAT from membranes into the cytosol. Internalized PI-PLC increased the amount of ChAT found in the cytosol, while reducing the amount of D-ChAT associated with membranes. In some tissues, endogenous glycosyl-phosphatidylinositol-specific phospholipase Cs (GPI-PLCs) function to release GPI-anchored proteins from membranes. To determine if an endogenous GPI-PLC might function to release D-ChAT from membranes, I utilized the GPI-PLC inhibitor zinc. Zinc inhibited an endogenous temperature-dependent release of ChAT from rat hippocampal minces, an endogenous release of ChAT from intracellular membranes into the cytosol of synaptosomes, and an endogenous conversion of ChAT from a detergent into an aqueous form in a plasma membrane enriched subcellular fraction (P4). These results suggest that some of the D-ChAT in rat hippocampal nerve terminals is GPI-anchored intracellularly, and that an endogenous GPI-PLC-like enzyme releases it from membranes into the cytosol. | |
| dc.format.mimetype | application/pdf | |
| dc.identifier.uri | http://hdl.handle.net/2346/14649 | en_US |
| dc.language.iso | eng | |
| dc.publisher | Texas Tech University | en_US |
| dc.rights.availability | Unrestricted. | |
| dc.subject | Neurotransmitters | en_US |
| dc.subject | Acetyltransferases | en_US |
| dc.subject | Hippocampus (Brain) | en_US |
| dc.subject | Membrane proteins | en_US |
| dc.title | Membrane binding characteristics of choline-o-acetyltransferase in rat hippocampal nerve terminals | |
| dc.type | Dissertation | |
| thesis.degree.department | TTUHSC -- Pharmacology and Neuroscience | |
| thesis.degree.discipline | TTUHSC -- Pharmacology and Neuroscience | |
| thesis.degree.grantor | Texas Tech University | |
| thesis.degree.level | Doctoral | |
| thesis.degree.name | Ph.D. |
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