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Purification and characterization of two protein kinases acting on the aquaporin SoPIP2; 1

Sjövall Larsen, Sara LU ; Alexandersson, Erik LU ; Johansson, Ingela LU ; Karlsson, Maria LU ; Johanson, Urban LU and Kjellbom, Per LU (2006) In Biochimica et Biophysica Acta - Biomembranes 1758(8). p.1157-1164
Abstract
Aquaporins are water channel proteins that facilitate the movement of water and other small solutes across biological membranes. Plants usually have large aquaporin families, providing them with many ways to regulate the water transport. Some aquaporins are regulated post-translationally by phosphorylation. We have previously shown that the water channel activity of SoP1P2;1, an aquaporin in the plasma membrane of spinach leaves, was enhanced by phosphorylation at Ser115 and Ser274. These two serine residues are highly conserved in all plasma membrane aquaporins of the PIP2 subgroup. In this study we have purified and characterized two protein kinases phosphorylating Ser115 and Ser274 in SoPIP2;1. By anion exchange chromatography, the... (More)
Aquaporins are water channel proteins that facilitate the movement of water and other small solutes across biological membranes. Plants usually have large aquaporin families, providing them with many ways to regulate the water transport. Some aquaporins are regulated post-translationally by phosphorylation. We have previously shown that the water channel activity of SoP1P2;1, an aquaporin in the plasma membrane of spinach leaves, was enhanced by phosphorylation at Ser115 and Ser274. These two serine residues are highly conserved in all plasma membrane aquaporins of the PIP2 subgroup. In this study we have purified and characterized two protein kinases phosphorylating Ser115 and Ser274 in SoPIP2;1. By anion exchange chromatography, the Ser115 kinase was purified from the soluble protein fraction isolated from spinach leaves. The Ca2+-dependent Ser274 kinase was purified by peptide affinity chromatography using plasma membranes isolated from spinach leaves. When characterized, the Ser115 kinase was Mg2+-dependent, Ca2+-independent and had a pH-optimum at 6.5. In accordance with previous studies using the oocyte expression system, site-directed mutagenesis and kinase and phosphatase inhibitors, the phosphorylation of Ser274, but not of Ser115, was increased in the presence of phosphatase inhibitors while kinase inhibitors decreased the phosphorylation of both Ser274 and Ser115. The molecular weight of the Ser274 kinase was approximately 50 kDa. The identification and characterization of these two protein kinases is an important step towards elucidating the signal transduction pathway for gating of the aquaporin SoPIP2;1. (Less)
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author
organization
publishing date
type
Contribution to journal
publication status
published
subject
keywords
protein kinase, phosphorylation, 1, SoPIP2, aquaporm, PIP2
in
Biochimica et Biophysica Acta - Biomembranes
volume
1758
issue
8
pages
1157 - 1164
publisher
Elsevier
external identifiers
  • wos:000241272900024
  • scopus:33748531849
ISSN
0005-2736
DOI
10.1016/j.bbamem.2006.06.002
language
English
LU publication?
yes
id
e2bf53ab-cd3a-4283-b984-caa185640c35 (old id 388223)
date added to LUP
2007-10-19 10:57:40
date last changed
2019-05-28 02:23:41
@article{e2bf53ab-cd3a-4283-b984-caa185640c35,
  abstract     = {Aquaporins are water channel proteins that facilitate the movement of water and other small solutes across biological membranes. Plants usually have large aquaporin families, providing them with many ways to regulate the water transport. Some aquaporins are regulated post-translationally by phosphorylation. We have previously shown that the water channel activity of SoP1P2;1, an aquaporin in the plasma membrane of spinach leaves, was enhanced by phosphorylation at Ser115 and Ser274. These two serine residues are highly conserved in all plasma membrane aquaporins of the PIP2 subgroup. In this study we have purified and characterized two protein kinases phosphorylating Ser115 and Ser274 in SoPIP2;1. By anion exchange chromatography, the Ser115 kinase was purified from the soluble protein fraction isolated from spinach leaves. The Ca2+-dependent Ser274 kinase was purified by peptide affinity chromatography using plasma membranes isolated from spinach leaves. When characterized, the Ser115 kinase was Mg2+-dependent, Ca2+-independent and had a pH-optimum at 6.5. In accordance with previous studies using the oocyte expression system, site-directed mutagenesis and kinase and phosphatase inhibitors, the phosphorylation of Ser274, but not of Ser115, was increased in the presence of phosphatase inhibitors while kinase inhibitors decreased the phosphorylation of both Ser274 and Ser115. The molecular weight of the Ser274 kinase was approximately 50 kDa. The identification and characterization of these two protein kinases is an important step towards elucidating the signal transduction pathway for gating of the aquaporin SoPIP2;1.},
  author       = {Sjövall Larsen, Sara and Alexandersson, Erik and Johansson, Ingela and Karlsson, Maria and Johanson, Urban and Kjellbom, Per},
  issn         = {0005-2736},
  keyword      = {protein kinase,phosphorylation,1,SoPIP2,aquaporm,PIP2},
  language     = {eng},
  number       = {8},
  pages        = {1157--1164},
  publisher    = {Elsevier},
  series       = {Biochimica et Biophysica Acta - Biomembranes},
  title        = {Purification and characterization of two protein kinases acting on the aquaporin SoPIP2; 1},
  url          = {http://dx.doi.org/10.1016/j.bbamem.2006.06.002},
  volume       = {1758},
  year         = {2006},
}