Characterization of deltoid, a chimeric protein containing the oligomerization site of hepatitis delta antigen.
Both forms of the hepatitis delta antigen (HDAg) encoded by hepatitis delta virus are active only as oligomers. Previous studies showed that quadrin, a synthetic 50-residue peptide containing residues 12-60 from the N-terminus of HDAg, interferes with HDAg oligomerization, forms an alpha-helical coiled coil in solution, and forms a novel square octamer in the crystal consisting of four antiparallel coiled-coil dimers joined at the corners by hydrophobic binding of oligomerization sites located at each end of the dimers. We designed and synthesized deltoid (CH3CO-[Cys23]HDAg-(12-27)-seryl-tRNA synthetae-(59-65)-[Cys42]HDAg-(34-60)-Tyr-NH2), a chimeric protein that structurally resembles one end of the quadrin dimer and contains a single oligomerization site. The 51-residue chain of deltoid contains a seven-residue alpha-hairpin loop in place of the remainder of the quadrin dimer plus Cys12 and Cys31 for forming an intrachain disulfide bridge. Reduced, unbridged deltoid (Tm=61 degrees C, DeltaG(H2O)=-1.7 kcal mol(-1)) was less stable to denaturation by heat or guanidine HCl than oxidized, intrachain disulfide-bridged deltoid (Tm>80 degrees C, DeltaG(H2O)=-2.6 kcal mol(-1)). Each form is an alpha-helical dimer that reversibly dissociates into two monomers (Kd=80 microM).
Saderholm
Matthew J
MJ
Department of Chemistry, The University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA. matthew_saderholm@berea.edu
Lemon
Stanley M
SM
Erickson
Bruce W
BW
D000595:Amino Acid Sequence; D002942:Circular Dichroism; D019281:Dimerization; D019791:Guanidine / Q000494:pharmacology; D006508:Hepatitis Antibodies / Q000276:immunology; D035923:Hepatitis delta Antigens / Q000737:chemistry* / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D019906:Nuclear Magnetic Resonance, Biomolecular; D011489:Protein Denaturation / Q000187:drug effects; D017510:Protein Folding; D020836:Protein Structure, Quaternary; D017433:Protein Structure, Secondary; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D013696:Temperature; D017186:Titrimetry
D006508:Hepatitis Antibodies; D035923:Hepatitis delta Antigens; D011993:Recombinant Fusion Proteins; D019791:Guanidine