Minisymposium 25: Protein Targeting
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Abs #
M2501: The Membrane Occupation Repeat Nexus (MORN) motifs in Arabidopsis PtdInsP Kinase 1 affect enzyme activity and subcellular localization
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Presenter: |
Im, Yang Ju Contact Presenter |
Authors | Im, Yang Ju (A) Davis, Amanda J (A) Eva, Johannes (A) Imara, Perera (A) Allen, Nina S (A) Boss, Wendy (A) | | Affiliations: |
(A): North Carolina State University
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The type I B family of phosphatidylinositol phosphate kinases (PIPKs) from Arabidopsis is characterized by a region of N-terminal putative MORN motifs. These motifs are not found in PIPKs from other eukaryotes. In theory, these MORN motifs should bind to membrane lipids and affect the subcellular distribution of the enzyme. We have used in vitro and in vivo studies to characterize the function of the MORN motifs of AtPIPK1. Recombinant truncated and full length proteins were characterized for enzyme activity. Deletion of the N-terminal MORN region (amino acid #1-252) increased the activity of the remaining truncated C-terminal catalytic domain (#253-752) >4 fold. When the MORN region was added back to the truncated catalytic domain, it enhanced the Vmax of the truncated catalytic domain but it had no effect on activity of the full-length enzyme. Lipid binding studies indicated that the MORN region bound to both the substrate, PtdIns(4)P, and the product, PtdIns(4,5)P2. These data suggested that the MORN region only increased enzyme activity if the active site was accessible. Our current model is that the MORN region acts as inhibitory regulation of the enzyme and protects the active site of the lipid kinase until the enzyme associates with a lipid bilayer; then the catalytic domain opens up and the MORN delivers PtdIns(4)P and removes PtdIns(4,5)P2. When expressed in tobacco cells the GFP- MORN region is recovered in the plasma membrane fraction. However, if the cells are osmotically stressed prior to membrane isolation, the GFP-MORN is found primarily in the lower phase suggesting that the MORN region is important for membrane binding and subcellular localization. This work was supported by a grant from the NSF (WFB).
