Evidence of intramolecular regulation of the Dictyostelium discoideum 34 000 Da F-actin-bundling protein.
Intramolecular interaction within the Ca(2+)-regulated 34 kDa actin-bundling protein from Dictyostelium discoideum was found to contribute to the regulation of its actin-binding activity. Recombinant N-terminally truncated proteins aa77-295, 124-295, and 139-295 bound actin at > or = 2:1 stoichiometry, which is 5-fold greater than the intact protein aa1-295 as assessed by cosedimentation with F-actin. These proteins also have enhanced cross-linking activity as assessed by viscometry and electron microscopy. All truncated 34 kDa proteins failed to bind (45)Ca(2+) on blots and displayed Ca(2+)-insensitive binding with actin, although most proteins possessed intact putative EF-hand Ca(2+)-binding motifs. An intramolecular interaction within the 34 kDa protein was inferred from direct demonstrations of domain-domain interaction among the truncated 34 kDa proteins both in the presence and absence of actin. The intramolecular interaction between interaction zone 1 (aa71-123) and interaction zone 2 (aa193-254) is proposed to maintain the N-terminal inhibitory region (aa1-76) in close proximity with the strong actin-binding site (aa193-254) in order to modulate the interaction of the intact protein with actin filaments.
Lim
R W
RW
Department of Cellular Biology, University of Georgia, Athens 30602, USA.
Furukawa
R
R
Fechheimer
M
M
D000199:Actins / Q000378:metabolism*; D000818:Animals; D001665:Binding Sites / Q000235:genetics; D002118:Calcium / Q000378:metabolism; D002352:Carrier Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D004023:Dictyostelium / Q000378:metabolism*; D008840:Microfilament Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008970:Molecular Weight; D010446:Peptide Fragments / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding / Q000235:genetics; D017434:Protein Structure, Tertiary / Q000235:genetics; D011817:Rabbits; D011994:Recombinant Proteins / Q000378:metabolism; D017384:Sequence Deletion
D000199:Actins; D002352:Carrier Proteins; C034037:F-actin-binding proteins; D008840:Microfilament Proteins; D010446:Peptide Fragments; D011994:Recombinant Proteins; C052523:actin filament bundling proteins; D002118:Calcium