Thymosin beta 4 binds actin in an extended conformation and contacts both the barbed and pointed ends.
The beta-thymosins are a family of highly polar peptides which serve in vivo to maintain a reservoir of unpolymerized actin monomers. In vitro, beta-thymosins form 1:1 complexes with actin monomers and inhibit both polymerization and exchange of the bound nucleotide. Circular dichroism data indicate that free thymosin beta 4 is predominantly unstructured, containing at most six residues of alpha-helix, and that up to six additional residues may adopt an alpha-helical conformation upon binding actin. NMR data indicate that many parts of thymosin beta 4 are not in tight contact with actin. Contacts between specific residues in actin and thymosin beta 4 were identified by zero-length cross-linking followed by isolation and sequencing of cross-linked peptides. After carbodiimide-mediated cross-linking, Lys-3 of thymosin beta 4 was cross-linked to Glu-167 of actin, and Lys-18 of thymosin beta 4 was cross-linked to one of the the N-terminal acidic residues of actin (Asp-1-Glu-4); the cross-linked actin residues lie within subdomains 3 and 1, respectively. These two contacts flank the alpha-helical region of thymosin beta 4 and place it on the barbed end; thymosin beta 4 can thus block actin polymerization sterically. After transglutaminase-mediated cross-linking, Lys-38 of thymosin beta 4 was cross-linked to Gln-41 of actin, placing the C-terminal region of thymosin beta 4 in contact with subdomain 2 on the pointed end; thymosin beta 4 may sterically block actin polymerization at the pointed end as well as the barbed end of the monomer. The distance between the pointed-end and barbed-end contacts requires that the C-terminal half of thymosin beta 4 be in a predominantly extended conformation.
Safer
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Department of Cell and Developmental Biology, University of Pennsylvania, Philadelphia 19104-6058, USA.
Sosnick
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TR
Elzinga
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D000199:Actins / Q000378:metabolism*; D000818:Animals; D002234:Carbodiimides; D002384:Catalysis; D002942:Circular Dichroism; D003432:Cross-Linking Reagents; D008958:Models, Molecular; D010455:Peptides / Q000737:chemistry / Q000302:isolation & purification; D011485:Protein Binding; D011487:Protein Conformation; D011817:Rabbits; D013947:Thymosin / Q000737:chemistry / Q000378:metabolism*; D011503:Transglutaminases / Q000378:metabolism; D000099201:Thymosin beta(4)
D000199:Actins; D002234:Carbodiimides; D003432:Cross-Linking Reagents; D010455:Peptides; D013947:Thymosin; D011503:Transglutaminases; D000099201:Thymosin beta(4)