HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

J. Biol. Chem., Vol. 281, Issue 42, 31696-31704, October 20, 2006

This Article Full Text Full Text (PDF) All Versions of this Article: 281/42/31696    most recent M604087200v1 Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Shridas, P. Articles by Waechter, C. J. Search for Related Content PubMed PubMed Citation Articles by Shridas, P. Articles by Waechter, C. J. Social Bookmarking                      What's this?

Human Dolichol Kinase, a Polytopic Endoplasmic Reticulum Membrane Protein with a Cytoplasmically Oriented CTP-binding Site^*

From the Department of Molecular and Cellular Biochemistry, University of Kentucky College of Medicine, Lexington, Kentucky 40536

Dolichol kinase (DK) catalyzes the CTP-dependent phosphorylation of dolichol in the biosynthesis de novo and possibly the recycling of dolichyl monophosphate in yeast and mammals. A cDNA clone from human brain encoding the mammalian homologue, hDKp, of the yeast enzyme has recently been identified. In this study hDK has been overexpressed in Chinese hamster ovary cells and shown to be a polytopic membrane protein localized in the endoplasmic reticulum with an N terminus extended into the lumen and a cytoplasmically oriented C terminus. A conserved sequence, DXXAXXXGXXXGX₈KKTXEG, found in several enzymes utilizing CTP as substrate including DKs, phytol kinases, and several CDP-diacylglycerol synthetases has been identified, and the possibility that it is part of the CTP-binding domain of hDKp has been investigated. Topological studies indicate that the loop between transmembrane domains (TMD) 11 and TMD12 of hDKp, containing the putative CTP binding domain, faces the cytoplasm. Deletion of the loop between TMD11-12, hDK(459-474), or mutation of selected conserved residues within the cytoplasmic loop results in either a partial or total loss of activity and significant reductions in the affinity for CTP. In addition, the SEC59 gene in the yeast DK mutant was sequenced, and a G420D substitution was found. Conversion of the corresponding residue Gly-443 in hDKp to aspartic acid resulted in inactivation of the mammalian enzyme. These results extend the information on the topological arrangement of hDKp and indicate that the cytoplasmic loop between TMDs 11-12, containing the critical conserved residues, lysine 470 and lysine 471 in the ⁴⁷⁰KKTXEG⁴⁷⁵ motif, is part of the CTP-binding site in hDK.

Received for publication, April 28, 2006 , and in revised form, August 8, 2006.

^* This work was supported by National Institutes of Health Grant GM36035 (to C. J. W.). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

¹ To whom correspondence should be addressed: Dept. of Molecular and Cellular Biochemistry, University of Kentucky College of Medicine, BBSRB, 741 S. Limestone St., Lexington, KY 40536. E-mail: waechte{at}uky.edu.

CiteULike    Complore    Connotea    Del.icio.us    Digg    Reddit    Technorati    What's this?

This article has been cited by other articles:

				G.-S. Han, L. O'Hara, S. Siniossoglou, and G. M. Carman Characterization of the Yeast DGK1-encoded CTP-dependent Diacylglycerol Kinase J. Biol. Chem., July 18, 2008; 283(29): 20443 - 20453. [Abstract] [Full Text] [PDF]

				G.-S. Han, L. O'Hara, G. M. Carman, and S. Siniossoglou An Unconventional Diacylglycerol Kinase That Regulates Phospholipid Synthesis and Nuclear Membrane Growth J. Biol. Chem., July 18, 2008; 283(29): 20433 - 20442. [Abstract] [Full Text] [PDF]