Factor XI apple domains and protein dimerization.
The coagulation protease zymogen factor (F)XI is a disulfide bond-linked homodimer, a configuration that is necessary for protein secretion and function. The non-catalytic portion of the FXI polypeptide contains four repeats called apple domains (A1-A4). It is clear that FXI A4 plays a key role in dimer formation, however, the importance of other apple domains to this process has not been examined. We prepared recombinant FXI molecules in which apple domains were exchanged with those of the structurally homologous monomeric protein prekallikrein (PK). As expected, FXI/PK chimeras containing FXI A4 are dimers, while those with PK A4 are monomers. FXI A4 contains cysteine at position 321 that forms the interchain disulfide bond, while Cys321 in PK is unavailable for interchain bond formation because it is paired with Cys326. FXI/PK chimeras containing PK A4 were modified by changing Cys326 to glycine, leaving Cys321 unpaired (PKA4-Gly326). FXI with a PK A4 domain is a monomer, however, introducing PKA4-Gly326 results in a disulfide bond-linked dimer. This indicates that dimer formation can occur in the absence of FXI A4. In proteins containing PKA4-Gly326, replacing FXI A3 with PK A3 partially interferes with dimer formation, while substitution of A2, or A2 and A3 prevents dimer formation. PKA4-Gly326 cannot induce the native PK molecule to dimerize. The data indicate that FXI A2 and A3 make contributions to dimer formation. As these domains are involved in activities that require dimeric protein, it seems reasonable that they stabilize this conformation.
Cheng
Q
Q
Department of Pathology, Vanderbilt University, Nashville, Tennessee 37232-6307, USA.
Sun
M-F
MF
Kravtsov
D V
DV
Aktimur
A
A
Gailani
D
D
D001665:Binding Sites; D002850:Chromatography, Gel; D003001:Cloning, Molecular; D019281:Dimerization; D004220:Disulfides; D004591:Electrophoresis, Polyacrylamide Gel; D005164:Factor IX / Q000737:chemistry* / Q000235:genetics; D006801:Humans; D011288:Prekallikrein / Q000737:chemistry / Q000235:genetics; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins
D004220:Disulfides; D011993:Recombinant Fusion Proteins; D005164:Factor IX; D011288:Prekallikrein