skip to main content
OSTI.GOV title logo U.S. Department of Energy
Office of Scientific and Technical Information

Title: High affinity binding proteins for the regulatory subunit of cAMP-dependent protein kinase: Cloning, characterization and expression of P150 and P75

Abstract

cAMP-dependent protein kinase II-B appears to be adapted for function in the mammalian central nervous system (CNS) via the properties of its regulatory subunit (RII-B). RII-B is selectively expressed in brain, tightly associated with cerebral cortex membranes and avidly complexed by the bovine brain calmodulin-binding protein designated P75. Complexes of RII-B and P75 polypeptides can be purified to near-homogeneity from either membrane or cytosolic fractions of brain homogenates, suggested that the binding protein plays a role in determining the subcellular localization and/or other CNS-specific properties of protein kinase II-B. In general, a single high-affinity RII-B binding protein is expressed in the brains of mammals, but the size of the protein varies (e.g., cow: 75 kDa; rat: 150 kDa). To investigate these non-abundant and CNS-enriched RII-B binding proteins, cDNAs for bovine brain P75 and rat brain P150, the homologue of P75, have been cloned and characterized. The cDNAs were retrieved from bovine and rat lambda gt11 expression libraries using {sup 32}P-RII-B as a functional probe. cDNA inserts subcloned into pIN-IA2 or pET-3b expression plasmids directed the production of partial P75 and P150 polypeptides in E. coli that exhibited RII-B binding activity. P150 cDNAs hybridize with 3 rat mRNAs (7.3, 5.0, 4.2more » kb) that are present in brain and lung, but not other tissues. These mRNAs are not detected in fetal brain, but are expressed during the period of post-natal synaptogenesis and in adult rats. The P75 cDNA hybridizes to a 6 kb bovine brain mRNA. Finally, 3'-deletion analysis demonstrated that the C-terminal 15-25 amino acids of P150 or P75 are essential for binding with RII-B.« less

Authors:
Publication Date:
Research Org.:
Yeshiva Univ., New York, NY (USA)
OSTI Identifier:
7152133
Resource Type:
Miscellaneous
Resource Relation:
Other Information: Thesis (Ph.D)
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; PHOSPHOTRANSFERASES; DNA-CLONING; AMINO ACIDS; BIOASSAY; BRAIN; COWS; EXPERIMENTAL DATA; GENES; HYBRIDIZATION; MESSENGER-RNA; PHOSPHORUS 32; PLASMIDS; RADIOASSAY; RATS; ANIMALS; BETA DECAY RADIOISOTOPES; BETA-MINUS DECAY RADIOISOTOPES; BODY; CARBOXYLIC ACIDS; CATTLE; CELL CONSTITUENTS; CENTRAL NERVOUS SYSTEM; CLONING; DATA; DAYS LIVING RADIOISOTOPES; DNA HYBRIDIZATION; DOMESTIC ANIMALS; ENZYMES; INFORMATION; ISOTOPES; LIGHT NUCLEI; MAMMALS; NERVOUS SYSTEM; NUCLEI; NUCLEIC ACIDS; NUMERICAL DATA; ODD-ODD NUCLEI; ORGANIC ACIDS; ORGANIC COMPOUNDS; ORGANS; PHOSPHORUS ISOTOPES; PHOSPHORUS-GROUP TRANSFERASES; RADIOISOTOPES; RNA; RODENTS; RUMINANTS; TRANSFERASES; VERTEBRATES; 550201* - Biochemistry- Tracer Techniques

Citation Formats

Bregman, D B. High affinity binding proteins for the regulatory subunit of cAMP-dependent protein kinase: Cloning, characterization and expression of P150 and P75. United States: N. p., 1989. Web.
Bregman, D B. High affinity binding proteins for the regulatory subunit of cAMP-dependent protein kinase: Cloning, characterization and expression of P150 and P75. United States.
Bregman, D B. 1989. "High affinity binding proteins for the regulatory subunit of cAMP-dependent protein kinase: Cloning, characterization and expression of P150 and P75". United States.
@article{osti_7152133,
title = {High affinity binding proteins for the regulatory subunit of cAMP-dependent protein kinase: Cloning, characterization and expression of P150 and P75},
author = {Bregman, D B},
abstractNote = {cAMP-dependent protein kinase II-B appears to be adapted for function in the mammalian central nervous system (CNS) via the properties of its regulatory subunit (RII-B). RII-B is selectively expressed in brain, tightly associated with cerebral cortex membranes and avidly complexed by the bovine brain calmodulin-binding protein designated P75. Complexes of RII-B and P75 polypeptides can be purified to near-homogeneity from either membrane or cytosolic fractions of brain homogenates, suggested that the binding protein plays a role in determining the subcellular localization and/or other CNS-specific properties of protein kinase II-B. In general, a single high-affinity RII-B binding protein is expressed in the brains of mammals, but the size of the protein varies (e.g., cow: 75 kDa; rat: 150 kDa). To investigate these non-abundant and CNS-enriched RII-B binding proteins, cDNAs for bovine brain P75 and rat brain P150, the homologue of P75, have been cloned and characterized. The cDNAs were retrieved from bovine and rat lambda gt11 expression libraries using {sup 32}P-RII-B as a functional probe. cDNA inserts subcloned into pIN-IA2 or pET-3b expression plasmids directed the production of partial P75 and P150 polypeptides in E. coli that exhibited RII-B binding activity. P150 cDNAs hybridize with 3 rat mRNAs (7.3, 5.0, 4.2 kb) that are present in brain and lung, but not other tissues. These mRNAs are not detected in fetal brain, but are expressed during the period of post-natal synaptogenesis and in adult rats. The P75 cDNA hybridizes to a 6 kb bovine brain mRNA. Finally, 3'-deletion analysis demonstrated that the C-terminal 15-25 amino acids of P150 or P75 are essential for binding with RII-B.},
doi = {},
url = {https://www.osti.gov/biblio/7152133}, journal = {},
number = ,
volume = ,
place = {United States},
year = {Sun Jan 01 00:00:00 EST 1989},
month = {Sun Jan 01 00:00:00 EST 1989}
}

Miscellaneous:
Other availability
Please see Document Availability for additional information on obtaining the full-text document. Library patrons may search WorldCat to identify libraries that may hold this item.

Save / Share: