Dimerization/docking domain of the type Iα regulatory subunit of cAMP- dependent protein kinase: Requirements for dimerization and docking are distinct but overlapping

Poopak Banky, Lily Jun Shen Huang, Susan S. Taylor

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Abstract

Based on increasing evidence that the type I R subunits as well as the type II R subunits localize to specific subcellular sites, we have carried out an extensive characterization of the stable dimerization domain at the N terminus of RIα. Deletion mutants as well as alanine scanning mutagenesis were used to delineate critical regions as well as particular amino acids that are required for homodimerization. A set of nested deletion mutants defined a minimum core required for dimerization. Two single site mutations on the C37H template, RIα(F47A) and RIα(F52A), were sufficient to abolish dimerization. In addition to serving as a dimerization motif, this domain also serves as a docking surface for binding to dual specificity anchoring proteins (D-AKAPs) (Huang, L. J., Durick, K., Weiner, J. A., Chun, J., and Taylor, S.S. (1997) J. Biol. Chem. 272, 8057-8064; Huang, L. J., Durick, K., Weiner, J. A., Chun, J., and Taylor, S.S. (1997) Proc. Natl. Acad. Sci. U.S.A. 94, 11184-11189). A similar strategy was used to map the sequence requirements for anchoring of RIα to D-AKAP1. Although dimerization appears to be essential for anchoring to D-AKAP1, anchoring can also be abolished by the following single site mutations: C37H, V20A, and I25A. These sites define 'hot spots' for the anchoring surface since each of these dimeric proteins are deficient in binding to D-AKAP1. In contrast to earlier predictions, the alignment of the dimerization/docking domains of RIα and RII show striking similarities yet subtle differences not only in their secondary structure (Newlon, M. G., Roy, M., Hausken, Z. E., Scott, J. D., and Jennings. P. A. (1997) J. Biol. Chem. 272, 23637-23644) but also in the distribution of residues important for both docking and dimerization functions.

Original languageEnglish (US)
Pages (from-to)35048-35055
Number of pages8
JournalJournal of Biological Chemistry
Volume273
Issue number52
DOIs
StatePublished - Dec 25 1998

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Dimerization
Cyclic AMP-Dependent Protein Kinases
Mutagenesis
Mutation
Alanine
Proteins
Scanning
Amino Acids

ASJC Scopus subject areas

  • Biochemistry

Cite this

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title = "Dimerization/docking domain of the type Iα regulatory subunit of cAMP- dependent protein kinase: Requirements for dimerization and docking are distinct but overlapping",
abstract = "Based on increasing evidence that the type I R subunits as well as the type II R subunits localize to specific subcellular sites, we have carried out an extensive characterization of the stable dimerization domain at the N terminus of RIα. Deletion mutants as well as alanine scanning mutagenesis were used to delineate critical regions as well as particular amino acids that are required for homodimerization. A set of nested deletion mutants defined a minimum core required for dimerization. Two single site mutations on the C37H template, RIα(F47A) and RIα(F52A), were sufficient to abolish dimerization. In addition to serving as a dimerization motif, this domain also serves as a docking surface for binding to dual specificity anchoring proteins (D-AKAPs) (Huang, L. J., Durick, K., Weiner, J. A., Chun, J., and Taylor, S.S. (1997) J. Biol. Chem. 272, 8057-8064; Huang, L. J., Durick, K., Weiner, J. A., Chun, J., and Taylor, S.S. (1997) Proc. Natl. Acad. Sci. U.S.A. 94, 11184-11189). A similar strategy was used to map the sequence requirements for anchoring of RIα to D-AKAP1. Although dimerization appears to be essential for anchoring to D-AKAP1, anchoring can also be abolished by the following single site mutations: C37H, V20A, and I25A. These sites define 'hot spots' for the anchoring surface since each of these dimeric proteins are deficient in binding to D-AKAP1. In contrast to earlier predictions, the alignment of the dimerization/docking domains of RIα and RII show striking similarities yet subtle differences not only in their secondary structure (Newlon, M. G., Roy, M., Hausken, Z. E., Scott, J. D., and Jennings. P. A. (1997) J. Biol. Chem. 272, 23637-23644) but also in the distribution of residues important for both docking and dimerization functions.",
author = "Poopak Banky and Huang, {Lily Jun Shen} and Taylor, {Susan S.}",
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T1 - Dimerization/docking domain of the type Iα regulatory subunit of cAMP- dependent protein kinase

T2 - Requirements for dimerization and docking are distinct but overlapping

AU - Banky, Poopak

AU - Huang, Lily Jun Shen

AU - Taylor, Susan S.

PY - 1998/12/25

Y1 - 1998/12/25

N2 - Based on increasing evidence that the type I R subunits as well as the type II R subunits localize to specific subcellular sites, we have carried out an extensive characterization of the stable dimerization domain at the N terminus of RIα. Deletion mutants as well as alanine scanning mutagenesis were used to delineate critical regions as well as particular amino acids that are required for homodimerization. A set of nested deletion mutants defined a minimum core required for dimerization. Two single site mutations on the C37H template, RIα(F47A) and RIα(F52A), were sufficient to abolish dimerization. In addition to serving as a dimerization motif, this domain also serves as a docking surface for binding to dual specificity anchoring proteins (D-AKAPs) (Huang, L. J., Durick, K., Weiner, J. A., Chun, J., and Taylor, S.S. (1997) J. Biol. Chem. 272, 8057-8064; Huang, L. J., Durick, K., Weiner, J. A., Chun, J., and Taylor, S.S. (1997) Proc. Natl. Acad. Sci. U.S.A. 94, 11184-11189). A similar strategy was used to map the sequence requirements for anchoring of RIα to D-AKAP1. Although dimerization appears to be essential for anchoring to D-AKAP1, anchoring can also be abolished by the following single site mutations: C37H, V20A, and I25A. These sites define 'hot spots' for the anchoring surface since each of these dimeric proteins are deficient in binding to D-AKAP1. In contrast to earlier predictions, the alignment of the dimerization/docking domains of RIα and RII show striking similarities yet subtle differences not only in their secondary structure (Newlon, M. G., Roy, M., Hausken, Z. E., Scott, J. D., and Jennings. P. A. (1997) J. Biol. Chem. 272, 23637-23644) but also in the distribution of residues important for both docking and dimerization functions.

AB - Based on increasing evidence that the type I R subunits as well as the type II R subunits localize to specific subcellular sites, we have carried out an extensive characterization of the stable dimerization domain at the N terminus of RIα. Deletion mutants as well as alanine scanning mutagenesis were used to delineate critical regions as well as particular amino acids that are required for homodimerization. A set of nested deletion mutants defined a minimum core required for dimerization. Two single site mutations on the C37H template, RIα(F47A) and RIα(F52A), were sufficient to abolish dimerization. In addition to serving as a dimerization motif, this domain also serves as a docking surface for binding to dual specificity anchoring proteins (D-AKAPs) (Huang, L. J., Durick, K., Weiner, J. A., Chun, J., and Taylor, S.S. (1997) J. Biol. Chem. 272, 8057-8064; Huang, L. J., Durick, K., Weiner, J. A., Chun, J., and Taylor, S.S. (1997) Proc. Natl. Acad. Sci. U.S.A. 94, 11184-11189). A similar strategy was used to map the sequence requirements for anchoring of RIα to D-AKAP1. Although dimerization appears to be essential for anchoring to D-AKAP1, anchoring can also be abolished by the following single site mutations: C37H, V20A, and I25A. These sites define 'hot spots' for the anchoring surface since each of these dimeric proteins are deficient in binding to D-AKAP1. In contrast to earlier predictions, the alignment of the dimerization/docking domains of RIα and RII show striking similarities yet subtle differences not only in their secondary structure (Newlon, M. G., Roy, M., Hausken, Z. E., Scott, J. D., and Jennings. P. A. (1997) J. Biol. Chem. 272, 23637-23644) but also in the distribution of residues important for both docking and dimerization functions.

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