Characterization of the substrate specificity of sucrose-phosphate synthase protein kinase

Robert W. McMichael, Jan Kochansky, Robert R. Klein, Steven C. Huber

Research output: Contribution to journalArticle

Abstract

Sucrose-phosphate synthase (SPS; EC 2.4.1.14) is regulated by reversible protein phosphorylation. When the enzyme is phosphorylated it is inactivated and can be reactivated by removal of phosphate. The major regulatory phosphorylation site is known to be Ser158 in the spinach-leaf enzyme, and two protein kinase activities have been resolved chromatographically which phosphorylate SPS at this site in vitro. In this report, we use a set of synthetic peptide substrate analogs based on the phosphorylation site sequence, and a set of Escherichia coli-expressed 26-kDa fragments of spinach SPS which contain the site, to identify the recognition elements that target the two protein kinases to Ser158. The major recognition element consists of basic residues at P-3 and P-6 relative to the phosphorylated serine. Comparison of the spinach enzyme amino-acid sequence with two other plant species show conservation of these amino acids and implies that these signals are also conserved. We also present evidence that glucose-6-phosphate is not only an allosteric activator of SPS but also an inhibitor of SPS-protein kinase per se, thereby allowing it to act at both levels of SPS regulation.

Original languageEnglish (US)
Pages (from-to)71-75
Number of pages5
JournalArchives of Biochemistry and Biophysics
Volume321
Issue number1
DOIs
StatePublished - Jan 1 1995

Fingerprint

sucrose-phosphate synthase
Phosphorylation
Spinacia oleracea
Substrate Specificity
Protein Kinases
Substrates
Enzymes
Amino Acids
Glucose-6-Phosphate
Protein Kinase Inhibitors
Serine
Escherichia coli
Amino Acid Sequence
Conservation
Phosphates
Peptides
Proteins

Keywords

  • Protein kinase
  • Recognition elements
  • Regulatory phosphorylation
  • Synthetic peptides

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Molecular Biology

Cite this

Characterization of the substrate specificity of sucrose-phosphate synthase protein kinase. / McMichael, Robert W.; Kochansky, Jan; Klein, Robert R.; Huber, Steven C.

In: Archives of Biochemistry and Biophysics, Vol. 321, No. 1, 01.01.1995, p. 71-75.

Research output: Contribution to journalArticle

McMichael, Robert W. ; Kochansky, Jan ; Klein, Robert R. ; Huber, Steven C. / Characterization of the substrate specificity of sucrose-phosphate synthase protein kinase. In: Archives of Biochemistry and Biophysics. 1995 ; Vol. 321, No. 1. pp. 71-75.
@article{2ff976f6d9444dceb9dba6936e104a82,
title = "Characterization of the substrate specificity of sucrose-phosphate synthase protein kinase",
abstract = "Sucrose-phosphate synthase (SPS; EC 2.4.1.14) is regulated by reversible protein phosphorylation. When the enzyme is phosphorylated it is inactivated and can be reactivated by removal of phosphate. The major regulatory phosphorylation site is known to be Ser158 in the spinach-leaf enzyme, and two protein kinase activities have been resolved chromatographically which phosphorylate SPS at this site in vitro. In this report, we use a set of synthetic peptide substrate analogs based on the phosphorylation site sequence, and a set of Escherichia coli-expressed 26-kDa fragments of spinach SPS which contain the site, to identify the recognition elements that target the two protein kinases to Ser158. The major recognition element consists of basic residues at P-3 and P-6 relative to the phosphorylated serine. Comparison of the spinach enzyme amino-acid sequence with two other plant species show conservation of these amino acids and implies that these signals are also conserved. We also present evidence that glucose-6-phosphate is not only an allosteric activator of SPS but also an inhibitor of SPS-protein kinase per se, thereby allowing it to act at both levels of SPS regulation.",
keywords = "Protein kinase, Recognition elements, Regulatory phosphorylation, Synthetic peptides",
author = "McMichael, {Robert W.} and Jan Kochansky and Klein, {Robert R.} and Huber, {Steven C.}",
year = "1995",
month = "1",
day = "1",
doi = "10.1006/abbi.1995.1369",
language = "English (US)",
volume = "321",
pages = "71--75",
journal = "Archives of Biochemistry and Biophysics",
issn = "0003-9861",
publisher = "Academic Press Inc.",
number = "1",

}

TY - JOUR

T1 - Characterization of the substrate specificity of sucrose-phosphate synthase protein kinase

AU - McMichael, Robert W.

AU - Kochansky, Jan

AU - Klein, Robert R.

AU - Huber, Steven C.

PY - 1995/1/1

Y1 - 1995/1/1

N2 - Sucrose-phosphate synthase (SPS; EC 2.4.1.14) is regulated by reversible protein phosphorylation. When the enzyme is phosphorylated it is inactivated and can be reactivated by removal of phosphate. The major regulatory phosphorylation site is known to be Ser158 in the spinach-leaf enzyme, and two protein kinase activities have been resolved chromatographically which phosphorylate SPS at this site in vitro. In this report, we use a set of synthetic peptide substrate analogs based on the phosphorylation site sequence, and a set of Escherichia coli-expressed 26-kDa fragments of spinach SPS which contain the site, to identify the recognition elements that target the two protein kinases to Ser158. The major recognition element consists of basic residues at P-3 and P-6 relative to the phosphorylated serine. Comparison of the spinach enzyme amino-acid sequence with two other plant species show conservation of these amino acids and implies that these signals are also conserved. We also present evidence that glucose-6-phosphate is not only an allosteric activator of SPS but also an inhibitor of SPS-protein kinase per se, thereby allowing it to act at both levels of SPS regulation.

AB - Sucrose-phosphate synthase (SPS; EC 2.4.1.14) is regulated by reversible protein phosphorylation. When the enzyme is phosphorylated it is inactivated and can be reactivated by removal of phosphate. The major regulatory phosphorylation site is known to be Ser158 in the spinach-leaf enzyme, and two protein kinase activities have been resolved chromatographically which phosphorylate SPS at this site in vitro. In this report, we use a set of synthetic peptide substrate analogs based on the phosphorylation site sequence, and a set of Escherichia coli-expressed 26-kDa fragments of spinach SPS which contain the site, to identify the recognition elements that target the two protein kinases to Ser158. The major recognition element consists of basic residues at P-3 and P-6 relative to the phosphorylated serine. Comparison of the spinach enzyme amino-acid sequence with two other plant species show conservation of these amino acids and implies that these signals are also conserved. We also present evidence that glucose-6-phosphate is not only an allosteric activator of SPS but also an inhibitor of SPS-protein kinase per se, thereby allowing it to act at both levels of SPS regulation.

KW - Protein kinase

KW - Recognition elements

KW - Regulatory phosphorylation

KW - Synthetic peptides

UR - http://www.scopus.com/inward/record.url?scp=0029081381&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0029081381&partnerID=8YFLogxK

U2 - 10.1006/abbi.1995.1369

DO - 10.1006/abbi.1995.1369

M3 - Article

C2 - 7639538

AN - SCOPUS:0029081381

VL - 321

SP - 71

EP - 75

JO - Archives of Biochemistry and Biophysics

JF - Archives of Biochemistry and Biophysics

SN - 0003-9861

IS - 1

ER -