Structural conservation between the actin monomer-binding sites of twinfilin and actin-depolymerizing factor (ADF)/cofilin.
Twinfilin is an evolutionarily conserved actin monomer-binding protein that regulates cytoskeletal dynamics in organisms from yeast to mammals. It is composed of two actin-depolymerization factor homology (ADF-H) domains that show approximately 20% sequence identity to ADF/cofilin proteins. In contrast to ADF/cofilins, which bind both G-actin and F-actin and promote filament depolymerization, twinfilin interacts only with G-actin. To elucidate the molecular mechanisms of twinfilin-actin monomer interaction, we determined the crystal structure of the N-terminal ADF-H domain of twinfilin and mapped its actin-binding site by site-directed mutagenesis. This domain has similar overall structure to ADF/cofilins, and the regions important for actin monomer binding in ADF/cofilins are especially well conserved in twinfilin. Mutagenesis studies show that the N-terminal ADF-H domain of twinfilin and ADF/cofilins also interact with actin monomers through similar interfaces, although the binding surface is slightly extended in twinfilin. In contrast, the regions important for actin-filament interactions in ADF/cofilins are structurally different in twinfilin. This explains the differences in actin-interactions (monomer versus filament binding) between twinfilin and ADF/cofilins. Taken together, our data show that the ADF-H domain is a structurally conserved actin-binding motif and that relatively small structural differences at the actin interfaces of this domain are responsible for the functional variation between the different classes of ADF-H domain proteins.
Program in Cellular Biotechnology, Institute of Biotechnology, P.O. Box 56, University of Helsinki, 00014 Helsinki, Finland.
Ville O
VO
Paavilainen
Michael C
MC
Merckel
Sandra
S
Falck
Pauli J
PJ
Ojala
Ehmke
E
Pohl
Matthias
M
Wilmanns
Pekka
P
Lappalainen
D051339:Actin Depolymerizing Factors; D000199:Actins / Q000737:chemistry* / Q000378:metabolism*; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D001665:Binding Sites; D003001:Cloning, Molecular; D017124:Conserved Sequence; D018360:Crystallography, X-Ray; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D019143:Evolution, Molecular; D007700:Kinetics; D051379:Mice; D008840:Microfilament Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D017433:Protein Structure, Secondary; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid
D051339:Actin Depolymerizing Factors; D000199:Actins; D008840:Microfilament Proteins; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; C485190:Twf1 protein, mouse; C113980:TWF1 protein, S cerevisiae