Document Detail


Identification and characterization of the nuclear isoform of Drosophila melanogaster CTP:phosphocholine cytidylyltransferase.
MedLine Citation:
PMID:  18922025     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
CTP:phosphocholine cytidylyltransferase (CCT) catalyzes the conversion of phosphocholine and cytidine 5'-triphosphate (CTP) to CDP-choline for the eventual synthesis of phosphatidylcholine (PC). The enzyme is regulated by reversible association with cellular membranes, with the rate of catalysis increasing following membrane association. Two isoforms of CCT appear to be present in higher eukaryotes, including Drosophila melanogaster, which contains the tandem genes Cct1 and Cct2. Before this study, the CCT1 isoform had not been characterized and the cellular location of each enzyme was unknown. In this investigation, the cDNA encoding the CCT1 isoform from D. melanogaster has been cloned and the recombinant enzyme purified and characterized to determine catalytic properties and the effect of lipid vesicles on activity. CCT1 exhibited a V max of 23904 nmol of CDP-choline min (-1) mg (-1) and apparent K m values for phosphocholine and CTP of 2.29 and 1.21 mM, respectively, in the presence of 20 muM PC/oleate vesicles. Cytidylyltransferases require a divalent cation for catalysis, and the cation preference of CCT1 was found to be as follows: Mg (2+) > Mn (2+) = Co (2+) > Ca (2+) = Ni (2+) > Zn (2+). The activity of the enzyme is stimulated by a variety of lipids, including phosphatidylcholine, phosphatidylinositol, phosphatidylglycerol, phosphatidylserine, diphosphatidylglycerol, and the fatty acid oleate. Phosphatidylethanolamine and phosphatidic acid, however, did not have a significant effect on CCT1 activity. The cellular location of both CCT1 and CCT2 isoforms was elucidated by expressing green fluorescent fusion proteins in cultured D. melanogaster Schneider 2 cells. CCT1 was identified as the nuclear isoform, while CCT2 is cytoplasmic.
Authors:
Dana M Tilley; Chadrick R Evans; Troy M Larson; Kevin A Edwards; Jon A Friesen
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't; Research Support, U.S. Gov't, Non-P.H.S.     Date:  2008-10-16
Journal Detail:
Title:  Biochemistry     Volume:  47     ISSN:  1520-4995     ISO Abbreviation:  Biochemistry     Publication Date:  2008 Nov 
Date Detail:
Created Date:  2008-11-04     Completed Date:  2008-12-31     Revised Date:  -    
Medline Journal Info:
Nlm Unique ID:  0370623     Medline TA:  Biochemistry     Country:  United States    
Other Details:
Languages:  eng     Pagination:  11838-46     Citation Subset:  IM    
Affiliation:
Department of Chemistry, Illinois State University, Normal, Illinois 61790, USA.
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MeSH Terms
Descriptor/Qualifier:
Amino Acid Sequence
Animals
Cell Line
Cell Membrane / enzymology
Cell Nucleus / enzymology*
Choline-Phosphate Cytidylyltransferase / chemistry,  genetics,  metabolism*
Drosophila Proteins / chemistry,  genetics,  metabolism*
Isoenzymes / chemistry,  genetics,  metabolism
Models, Biological
Molecular Sequence Data
Sequence Homology, Amino Acid
Chemical
Reg. No./Substance:
0/Drosophila Proteins; 0/Isoenzymes; EC 2.7.7.15/Cct1protein, Drosophila; EC 2.7.7.15/Choline-Phosphate Cytidylyltransferase

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