| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
|
|
|
|||||||
|
|||||||||
J Biol Chem, Vol. 273, Issue 12, 6844-6852, March 20, 1998
,
From the In the yeast Saccharomyces
cerevisiae, choline kinase (ATP:choline phosphotransferase, EC
2.7.1.32) is the product of the CKI gene. Choline kinase
catalyzes the committed step in the synthesis of phosphatidylcholine by
the CDP-choline pathway. The yeast enzyme was overexpressed 106-fold in
Sf-9 insect cells and purified 71.2-fold to homogeneity from the
cytosolic fraction by chromatography with concanavalin A, Affi-Gel
Blue, and Mono Q. The N-terminal amino acid sequence of purified
choline kinase matched perfectly with the deduced sequence of the
CKI gene. The minimum subunit molecular mass (73 kDa) of
purified choline kinase was in good agreement with the predicted size
(66.3 kDa) of the CKI gene product. Native choline kinase
existed in oligomeric structures of dimers, tetramers, and octomers.
The amounts of the tetrameric and octomeric forms increased in the
presence of the substrate ATP. Antibodies were raised against the
purified enzyme and were used to identify choline kinase in insect
cells and in S. cerevisiae. Maximum choline kinase activity
was dependent on Mg2+ ions (10 mM) at pH 9.5 and at 30 °C. The equilibrium constant (0.2) for the reaction
indicated that the reverse reaction was favored in vitro.
The activation energy for the reaction was 6.26 kcal/mol, and the
enzyme was labile above 30 °C. Choline kinase exhibited saturation
kinetics with respect to choline and positive cooperative kinetics with
respect to ATP (n = 1.4-2.3). Results of the kinetic
experiments indicated that the enzyme catalyzes a sequential Bi
Bi reaction. The Vmax for the reaction was
138.7 µmol/min/mg, and the Km values for choline
and ATP were 0.27 mM and 90 µM, respectively.
The turnover number per choline kinase subunit was 153 s Department of Food Science, Cook College,
New Jersey Agricultural Experiment Station, Rutgers University, New
Brunswick, New Jersey 08903, the § Lord and Taylor
Laboratory for Lung Biochemistry and the Anna Perahia Adatto Clinical
Research Center, National Jewish Center for Immunology and Respiratory
Medicine, Denver, Colorado 80206, and the ¶ North American
Operations/Bioresearch, Beckman Instruments,
Somerset, New Jersey 08875
1. Ethanolamine was a poor substrate for the purified
choline kinase, and it was also poor inhibitor of choline kinase
activity. ADP inhibited choline kinase activity (IC50 = 0.32 mM) in a positive cooperative manner
(n = 1.5), and the mechanism of inhibition with
respect to ATP and choline was complex. The regulation of choline
kinase activity by ATP and ADP may be physiologically relevant.
This article has been cited by other articles:
|
|
 |
|
  A. Soto and G. M. Carman Regulation of the Saccharomyces cerevisiae CKI1-encoded Choline Kinase by Zinc Depletion J. Biol. Chem., April 11, 2008; 283(15): 10079 - 10088. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
G. M. Carman and S. A. Henry Phosphatidic Acid Plays a Central Role in the Transcriptional Regulation of Glycerophospholipid Synthesis in Saccharomyces cerevisiae J. Biol. Chem., December 28, 2007; 282(52): 37293 - 37297. [Full Text] [PDF]
|
|
|
|
|
|
M.-G. Choi, V. Kurnov, M. C. Kersting, A. Sreenivas, and G. M. Carman Phosphorylation of the Yeast Choline Kinase by Protein Kinase C: IDENTIFICATION OF Ser25 AND Ser30 AS MAJOR SITES OF PHOSPHORYLATION J. Biol. Chem., July 15, 2005; 280(28): 26105 - 26112. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. C. Kersting, H.-S. Choi, and G. M. Carman Regulation of the Yeast EKI1-encoded Ethanolamine Kinase by Inositol and Choline J. Biol. Chem., August 20, 2004; 279(34): 35353 - 35359. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 R. Roggero, R. Zufferey, M. Minca, E. Richier, M. Calas, H. Vial, and C. Ben Mamoun Unraveling the Mode of Action of the Antimalarial Choline Analog G25 in Plasmodium falciparum and Saccharomyces cerevisiae Antimicrob. Agents Chemother., August 1, 2004; 48(8): 2816 - 2824. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
C. Yuan and C. Kent Identification of Critical Residues of Choline Kinase A2 from Caenorhabditis elegans J. Biol. Chem., April 23, 2004; 279(17): 17801 - 17809. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. G. Howe, V. Zaremberg, and C. R. McMaster Cessation of Growth to Prevent Cell Death Due to Inhibition of Phosphatidylcholine Synthesis Is Impaired at 37 {degrees}C in Saccharomyces cerevisiae J. Biol. Chem., November 8, 2002; 277(46): 44100 - 44107. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
Y. Yu, A. Sreenivas, D. B. Ostrander, and G. M. Carman Phosphorylation of Saccharomyces cerevisiae Choline Kinase on Ser30 and Ser85 by Protein Kinase A Regulates Phosphatidylcholine Synthesis by the CDP-choline Pathway J. Biol. Chem., September 13, 2002; 277(38): 34978 - 34986. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. Li, J. J. Marsh, and R. G. Spragg Effect Of CTP:Phosphocholine Cytidylyltransferase Overexpression on the Mouse Lung Surfactant System Am. J. Respir. Cell Mol. Biol., June 1, 2002; 26(6): 709 - 715. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A. J. Ryan, D. M. McCoy, S. N. Mathur, F. J. Field, and R. K. Mallampalli Lipoprotein deprivation stimulates transcription of the CTP:phosphocholine cytidylyltransferase gene J. Lipid Res., August 1, 2000; 41(8): 1268 - 1277. [Abstract] [Full Text]
|
|
|
|
|
|
K. Kim, K.-H. Kim, M. K. Storey, D. R. Voelker, and G. M. Carman Isolation and Characterization of the Saccharomyces cerevisiae EKI1 Gene Encoding Ethanolamine Kinase J. Biol. Chem., May 21, 1999; 274(21): 14857 - 14866. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K.-H. Kim and G. M. Carman Phosphorylation and Regulation of Choline Kinase from Saccharomyces cerevisiae by Protein Kinase A J. Biol. Chem., April 2, 1999; 274(14): 9531 - 9538. [Abstract] [Full Text] [PDF]
|
|
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
| All ASBMB Journals | Molecular and Cellular Proteomics |
| Journal of Lipid Research | ASBMB Today |
