Serine phosphorylation of syndecan-2 proteoglycan cytoplasmic domain

Eok Soo Oh; John R. Couchman; Anne Woods

doi:10.1006/abbi.1997.0180

Serine phosphorylation of syndecan-2 proteoglycan cytoplasmic domain

Eok Soo Oh, John R. Couchman, Anne Woods

Department of Life Science

Research output: Contribution to journal › Article › peer-review

48 Scopus citations

Abstract

Protein kinase C (PKC) is involved in cell-matrix and cell-cell adhesion, and the cytoplasmic domain of syndecan-2 contains two serines (residues 197 and 198) which lie in a consensus sequence for phosphorylation by PKC. Other serine and threonine residues are present but not in a consensus sequence. We investigated phosphorylation of syndecan-2 cytoplasmic domain by PKC, using purified GST-syndecan-2 fusion proteins and synthetic peptides corresponding to regions of the cytoplasmic domain. A synthetic peptide encompassing the entire cytoplasmic domain of syndecan-2 was phosphorylated by PKC with high affinity. Peptide mapping and substitution studies showed that both serines were phosphoacceptors, but each had slightly different affinity, with that of serine-197 being higher than serine-198. The efficiency of phosphorylation was concentration-dependent. At low concentrations, the cytoplasmic domain peptides were monomeric, with 2 mol/mol serine phosphorylation. At higher concentrations, however, the peptides formed dimers, with only 0.5 mol/mol phosphorylation. Concentration- dependent dimerization was not altered by phosphorylation. Phosphorylation is, therefore, dependent on the conformation of syndecan-2 cytoplasmic domain, but does not affect its oligomeric status.

Original language	English
Pages (from-to)	67-74
Number of pages	8
Journal	Archives of Biochemistry and Biophysics
Volume	344
Issue number	1
DOIs	https://doi.org/10.1006/abbi.1997.0180
State	Published - 1 Aug 1997

Bibliographical note

Funding Information:
We thank Dr. J. T. Gallagher and his colleagues (University of Manchester, UK) for the provision of the full length and truncated syndecan-2 cDNA constructs. We also thank Dr. T. Lincoln (Department of Pathology, UAB) for help with PKA and PKG phosphorylation experiments. This work was supported in part by NIH Grant GM50194 to JRC, and by the Multipurpose Arthritis and Musculoskeletal Diseases Center at UAB, through NIH P60 Grant AR20614 to A.W.

Keywords

Heparan sulfate proteoglycan
Phosphorylation
Protein kinase C
Syndecan-2

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title = "Serine phosphorylation of syndecan-2 proteoglycan cytoplasmic domain",

abstract = "Protein kinase C (PKC) is involved in cell-matrix and cell-cell adhesion, and the cytoplasmic domain of syndecan-2 contains two serines (residues 197 and 198) which lie in a consensus sequence for phosphorylation by PKC. Other serine and threonine residues are present but not in a consensus sequence. We investigated phosphorylation of syndecan-2 cytoplasmic domain by PKC, using purified GST-syndecan-2 fusion proteins and synthetic peptides corresponding to regions of the cytoplasmic domain. A synthetic peptide encompassing the entire cytoplasmic domain of syndecan-2 was phosphorylated by PKC with high affinity. Peptide mapping and substitution studies showed that both serines were phosphoacceptors, but each had slightly different affinity, with that of serine-197 being higher than serine-198. The efficiency of phosphorylation was concentration-dependent. At low concentrations, the cytoplasmic domain peptides were monomeric, with 2 mol/mol serine phosphorylation. At higher concentrations, however, the peptides formed dimers, with only 0.5 mol/mol phosphorylation. Concentration- dependent dimerization was not altered by phosphorylation. Phosphorylation is, therefore, dependent on the conformation of syndecan-2 cytoplasmic domain, but does not affect its oligomeric status.",

keywords = "Heparan sulfate proteoglycan, Phosphorylation, Protein kinase C, Syndecan-2",

author = "Oh, {Eok Soo} and Couchman, {John R.} and Anne Woods",

note = "Funding Information: We thank Dr. J. T. Gallagher and his colleagues (University of Manchester, UK) for the provision of the full length and truncated syndecan-2 cDNA constructs. We also thank Dr. T. Lincoln (Department of Pathology, UAB) for help with PKA and PKG phosphorylation experiments. This work was supported in part by NIH Grant GM50194 to JRC, and by the Multipurpose Arthritis and Musculoskeletal Diseases Center at UAB, through NIH P60 Grant AR20614 to A.W.",

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N1 - Funding Information: We thank Dr. J. T. Gallagher and his colleagues (University of Manchester, UK) for the provision of the full length and truncated syndecan-2 cDNA constructs. We also thank Dr. T. Lincoln (Department of Pathology, UAB) for help with PKA and PKG phosphorylation experiments. This work was supported in part by NIH Grant GM50194 to JRC, and by the Multipurpose Arthritis and Musculoskeletal Diseases Center at UAB, through NIH P60 Grant AR20614 to A.W.

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N2 - Protein kinase C (PKC) is involved in cell-matrix and cell-cell adhesion, and the cytoplasmic domain of syndecan-2 contains two serines (residues 197 and 198) which lie in a consensus sequence for phosphorylation by PKC. Other serine and threonine residues are present but not in a consensus sequence. We investigated phosphorylation of syndecan-2 cytoplasmic domain by PKC, using purified GST-syndecan-2 fusion proteins and synthetic peptides corresponding to regions of the cytoplasmic domain. A synthetic peptide encompassing the entire cytoplasmic domain of syndecan-2 was phosphorylated by PKC with high affinity. Peptide mapping and substitution studies showed that both serines were phosphoacceptors, but each had slightly different affinity, with that of serine-197 being higher than serine-198. The efficiency of phosphorylation was concentration-dependent. At low concentrations, the cytoplasmic domain peptides were monomeric, with 2 mol/mol serine phosphorylation. At higher concentrations, however, the peptides formed dimers, with only 0.5 mol/mol phosphorylation. Concentration- dependent dimerization was not altered by phosphorylation. Phosphorylation is, therefore, dependent on the conformation of syndecan-2 cytoplasmic domain, but does not affect its oligomeric status.

AB - Protein kinase C (PKC) is involved in cell-matrix and cell-cell adhesion, and the cytoplasmic domain of syndecan-2 contains two serines (residues 197 and 198) which lie in a consensus sequence for phosphorylation by PKC. Other serine and threonine residues are present but not in a consensus sequence. We investigated phosphorylation of syndecan-2 cytoplasmic domain by PKC, using purified GST-syndecan-2 fusion proteins and synthetic peptides corresponding to regions of the cytoplasmic domain. A synthetic peptide encompassing the entire cytoplasmic domain of syndecan-2 was phosphorylated by PKC with high affinity. Peptide mapping and substitution studies showed that both serines were phosphoacceptors, but each had slightly different affinity, with that of serine-197 being higher than serine-198. The efficiency of phosphorylation was concentration-dependent. At low concentrations, the cytoplasmic domain peptides were monomeric, with 2 mol/mol serine phosphorylation. At higher concentrations, however, the peptides formed dimers, with only 0.5 mol/mol phosphorylation. Concentration- dependent dimerization was not altered by phosphorylation. Phosphorylation is, therefore, dependent on the conformation of syndecan-2 cytoplasmic domain, but does not affect its oligomeric status.

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Serine phosphorylation of syndecan-2 proteoglycan cytoplasmic domain

Abstract

Bibliographical note

Keywords

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