A joint project of the Graduate School, Peabody College, and the Jean & Alexander Heard Library

Title page for ETD etd-03192010-142021

Type of Document Dissertation
Author Zhou, Xiaoming
Author's Email Address xiaoming.zhou@vanderbilt.edu
URN etd-03192010-142021
Title Purification, characterization and reconstitution of Drs2 protein, a phospholipid flippase from budding yeast
Degree PhD
Department Biological Sciences
Advisory Committee
Advisor Name Title
Carl H. Johnson Committee Chair
Charles R. Sanders Committee Member
Hassane S. Mchaourab Committee Member
Katherine L. Friedman Committee Member
Todd R. Graham Committee Member
  • membrane asymmetry
  • P4-ATPase
  • proteoliposome
  • Cdc50
  • flippase
Date of Defense 2009-12-11
Availability unrestricted
Type-IV P-type ATPases are putative phospholipid flippases that translocate specific phospholipid substrates from the exofacial to the cytosolic leaflet of membranes to generate phospholipid asymmetry. However, flippase activity has not been reconstituted with any purified type-IV P-type ATPase, and so whether these ATPases directly pump phospholipid across the membrane bilayer is unknown. The major goal of this dissertation project is to test whether Drs2p, a type-IV P-type ATPase from yeast Saccharomyces cerevisiae, is a flippase or not. I show that Drs2p can directly catalyze phospholipid translocation through purification and reconstitution of this ATPase into artificial membranes. This flippase activity is specific to a phosphatidylserine analogue and requires active Drs2p. My data also suggest that only a portion of Drs2p molecules are active after purification and these active molecules may result from partial proteolysis within the carboxyl terminus of Drs2p. This observation is consistent with a model that the carboxyl tail is auto-inhibitory to Drs2p activity. In this study, I demonstrate for the first time the reconstitution of flippase activity with a purified type-IV P-type ATPase.
  Filename       Size       Approximate Download Time (Hours:Minutes:Seconds) 
 28.8 Modem   56K Modem   ISDN (64 Kb)   ISDN (128 Kb)   Higher-speed Access 
  Xiaoming_dissertation.pdf 3.89 Mb 00:18:00 00:09:15 00:08:06 00:04:03 00:00:20

Browse All Available ETDs by ( Author | Department )

If you have more questions or technical problems, please Contact LITS.