Primary structure and domain organization of human alpha and beta adducin.

Abstract

Abstract. Adducin is a membrane-skeletal protein which is a candidate to promote assembly of a spectrin-actin network in erythrocytes and at sites of cellcell contact in epithelial tissues . The complete sequence of both subunits of human adducin, alpha (737 amino acids), and beta (726 amino acids) has been deduced by analysis of the cDNAs . The two subunits have strikingly conserved amino acid sequences with 49% identity and 66% similarity, suggesting evolution by gene duplication . Each adducin subunit has three distinct domains: a 39-kD NH2-terminal globular protease-resistant domain, connected by a 9-kD domain to a 33-kD COOH-terminal protease-sensitive tail comprised almost entirely of hydrophilic amino acids . The tail is responsible for the high frictional ratio of adducin noted previously, and was visualized by EM . The head domains of both adducin subunits exhibit a limited sequence similarity with the NH2-terminal ac-LASMA membranes of eukaryotic cells contain a system of structural proteins known as the spectrin-based membrane skeleton that has a general role in organi zation ofcertain integral membrane proteins and of coupling these proteins to cytoplasmic proteins (2,16,29) . Spectrin, the principal component of the membrane skeleton, is a rodshaped protein that associates with F-actin ateach end . Spectrin and actin together with accessory proteins form a regular geodesic domelike structure in erythrocytes where short actin filaments are associated with five and seven spectrin molecules to form a polygonal network (5, 28) . The striking images of the spectrin skeleton raise the issue of how such a structure could be assembled and has focused attention on the accessory proteins that interact with spectrin and actin at spectrin-actin junctions (1) .Adducin is a membrane-skeletal protein that promotes association of spectrin with actin, and this interaction is regulated by calcium/calmodulin (14, 31) . Adducin is comprised of two subunits with M 103,000 (alpha) and 97,000 (beta) and was originally purified from human erythrocytes based on its calmodulin-binding activity (13) . Adducin also is a substrate for protein kinase C in intact cells as well as under in vitro conditions (6,27,33,39). A protein closely related

Headpiece domain of dematin is required for the stability of the erythrocyte membrane

“…Adducin, consisting of ␣ and ␤ subunits, promotes the association of spectrin with F-actin, bundles actin filaments, and caps barbed ends of actin filaments in vitro (31) (32)(33)(34). Surprisingly, recent generation of ␤ adducin null mice, which also lack Ϸ80% of ␣ adducin, revealed a nearly normal content of spectrin and actin in the erythrocyte ghosts (35,36).…”

Interaction of the SH2 Domain of Fyn with a Cytoskeletal Protein, β-Adducin

“…There are three members of adducin, ␣-, ␤-, ␥-adducin, which form a heteromeric protein complex comprised of either ␣/␤ subunits or ␣/␥ subunits (29 -32). ␣-and ␥-adducins are expressed in most tissues, whereas ␤-adducin is in more restricted tissues such as the brain and spleen (30,33). Because Fyn and ␤-adducin were abundant in the Nonidet P-40-insoluble fraction (Fig.…”

Role of the Gly460Trp polymorphism of the α-adducin gene in primary hypertension in Scandinavians

“…The primers were designed according to the published ␣-adducin cDNA sequence. 28 PCR was performed with 50 ng of genomic DNA in a total volume of 20 l containing 10 pmol of each primer, 2 nmol dNTPs, 0.5 U Taq polymerase (Perkin-Elmer, Foster City, CA, USA) in 1 × PCRbuffer for Taq-polymerase (10 mM Tris HCl pH 8.3; 50 mM KCl; 1.5 mM MgCl 2 ; 0.001% gelatin) (Perkin-Elmer), 1.5% formamide and 1.5 mM MgCl 2 . PCR conditions were as follows: initial denaturation at 94°C for 5 min, followed by 30 cycles of denaturation (94°C for 30 sec), annealing (60°C for 30 sec) and extension (72°C for 30 sec), with the final extension at 72°C for 10 min.…”