Cloning and expression of an intron-less gene for AKAP 75, an anchor protein for the regulatory subunit of cAMP-dependent protein kinase II beta.
The A-Kinase Anchor Protein AKAP 75 (formerly designated bovine brain P75) is a particulate brain protein that avidly binds the regulatory subunit (RII beta) of cAMP-dependent protein kinase II beta (Bregman, D. B., Hirsch, A.H. and Rubin, C.S. (1991) J. Biol. Chem. 266, 7207-7213). The formation of stable AKAP 75.RII beta complexes provides a potential mechanism for targeting physiological signals carried by cAMP to specific effector sites within neurons and other brain cells. We have now cloned and characterized the AKAP 75 gene. Its coding sequence is novel and unexpectedly short (1284 base pairs) and contains no introns. When the AKAP 75 gene was transfected into HEK 293 cells, a new RII beta-binding protein with an apparent Mr of 75,000 accumulated. A high proportion (approximately 65%) of the AKAP 75 gene product was excluded from the cytoplasm and was recovered in the 40,000 x g pellet derived from disrupted transfected cells. In contrast, cells transfected with a construct encoding 249 amino acids from the central and C-terminal regions of AKAP 75 produced an RII beta-binding protein (apparent Mr = 45,000) that was exclusively cytosolic. AKAP 75 is a novel protein composed of only 428 amino acid residues (Mr = 47,878). A highly acidic C-terminal region mediates the binding of RII beta (and cAMP-dependent protein kinase II beta), whereas a positively charged N-terminal segment contains structural features that are essential for the association of AKAP 75 with the cytoskeleton and/or intracellular membranes.
Hirsch
A H
AH
Department of Molecular Pharmacology, Albert Einstein College of Medicine, Bronx, New York 10461.
Glantz
S B
SB
Li
Y
Y
You
Y
Y
Rubin
C S
CS
D054758:A Kinase Anchor Proteins; D048868:Adaptor Proteins, Signal Transducing; D000595:Amino Acid Sequence; D000818:Animals; D001345:Autoradiography; D001483:Base Sequence; D015153:Blotting, Western; D002352:Carrier Proteins; D002417:Cattle; D002460:Cell Line; D003001:Cloning, Molecular; D004247:DNA / Q000235:genetics; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D007438:Introns; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins; D011494:Protein Kinases / Q000737:chemistry*; D011506:Proteins / Q000235:genetics*; D012689:Sequence Homology, Nucleic Acid
D054758:A Kinase Anchor Proteins; C484064:AKAP5 protein, Bos taurus; C487775:AKAP5 protein, human; D048868:Adaptor Proteins, Signal Transducing; D002352:Carrier Proteins; D009419:Nerve Tissue Proteins; D011506:Proteins; D004247:DNA; D011494:Protein Kinases