The C2 Domain of Phosphatidylserine Decarboxylase 2 Is Not Required for Catalysis but Is Essential for in VivoFunction
Open Access
- 1 September 2002
- journal article
- Published by Elsevier BV
- Vol. 277 (37), 33720-33726
- https://doi.org/10.1074/jbc.m205672200
Abstract
Phosphatidylserine decarboxylase 2 (Psd2p) is currently being used to study lipid trafficking processes in intact and permeabilized yeast cells. The Psd2p contains a C2 homology domain and a putative Golgi retention/localization (GR) domain. C2 domains play important functions in membrane binding and docking reactions involving phospholipids and proteins. We constructed a C2 domain deletion variant (C2Δ) and a GR deletion variant (GRΔ) of Psd2p and examined their effects on in vivo function and catalysis. Immunoblotting confirmed that the predicted immature and mature forms of Psd2(C2Δ)p, Psd2(GRΔ)p, and wild type Psd2p were produced in vivo and that the proteins localized normally. Enzymology revealed that the Psd2(C2Δ)p and Psd2(GRΔ)p were catalytically active and could readily be expressed at levels 10-fold higher than endogenous Psd2p. Both Psd2p and Psd2(GRΔ)p expression complemented the growth defect ofpsd1Δpsd2Δ strains and resulted in normal aminoglycerophospholipid metabolism. In contrast, the Psd2(C2Δ)p failed to complementpsd1Δpsd2Δ strains, and [3H]serine labeling revealed a severe defect in the formation of PtdEtn in both intact and permeabilized cells, indicative of disruption of lipid trafficking. These findings identify an essential, non-catalytic function of the C2 domain of Psd2p and raise the possibility that it plays a direct role in membrane docking and/or PtdSer transport to the enzyme.Keywords
This publication has 24 references indexed in Scilit:
- Membrane Targeting by C1 and C2 DomainsPublished by Elsevier BV ,2001
- Characterization of Phosphatidylserine Transport to the Locus of Phosphatidylserine Decarboxylase 2 in Permeabilized YeastJournal of Biological Chemistry, 2001
- A New Gene Involved in the Transport-dependent Metabolism of Phosphatidylserine, PSTB2/PDR17, Shares Sequence Similarity with the Gene Encoding the Phosphatidylinositol/Phosphatidylcholine Transfer Protein,SEC14Published by Elsevier BV ,2000
- The C2 Domains of Rabphilin3A Specifically Bind Phosphatidylinositol 4,5-Bisphosphate Containing Vesicles in a Ca2+-dependent MannerPublished by Elsevier BV ,1998
- Synthesis and Intracellular Transport of Aminoglycerophospholipids in Permeabilized Cells of the Yeast, Saccharomyces cerevisiaePublished by Elsevier BV ,1995
- Characterization of a microsomal subfraction associated with mitochondria of the yeast, Saccharomyces cerevisiae. Involvement in synthesis and import of phospholipids into mitochondriaBiochimica et Biophysica Acta (BBA) - Biomembranes, 1995
- Phosphatidylserine Decarboxylase 2 of Saccharomyces cerevisiáePublished by Elsevier BV ,1995
- Identification of a Non-mitochondrial Phosphatidylserine Decarboxylase Activity (PSD2) in the Yeast Saccharomyces cerevisiaeJournal of Biological Chemistry, 1995
- Site-directed mutagenesis by overlap extension using the polymerase chain reactionGene, 1989
- The Saccharomyces cerevisiae SEC14 gene encodes a cytosolic factor that is required for transport of secretory proteins from the yeast Golgi complex.The Journal of cell biology, 1989