Structure of Ca2+-bound S100A4 and its interaction with peptides derived from nonmuscle myosin-IIA.
S100A4, also known as mts1, is a member of the S100 family of Ca2+-binding proteins that is directly involved in tumor invasion and metastasis via interactions with specific protein targets, including nonmuscle myosin-IIA (MIIA). Human S100A4 binds two Ca2+ ions with the typical EF-hand exhibiting an affinity that is nearly 1 order of magnitude tighter than that of the pseudo-EF-hand. To examine how Ca2+ modifies the overall organization and structure of the protein, we determined the 1.7 A crystal structure of the human Ca2+-S100A4. Ca2+ binding induces a large reorientation of helix 3 in the typical EF-hand. This reorganization exposes a hydrophobic cleft that is comprised of residues from the hinge region,helix 3, and helix 4, which afford specific target recognition and binding. The Ca2+-dependent conformational change is required for S100A4 to bind peptide sequences derived from the C-terminal portion of the MIIA rod with submicromolar affinity. In addition, the level of binding of Ca2+ to both EF-hands increases by 1 order of magnitude in the presence of MIIA. NMR spectroscopy studies demonstrate that following titration with a MIIA peptide, the largest chemical shift perturbations and exchange broadening effects occur for residues in the hydrophobic pocket of Ca2+-S100A4. Most of these residues are not exposed in apo-S100A4 and explain the Ca2+ dependence of formation of theS100A4-MIIA complex. These studies provide the foundation for understanding S100A4 target recognition and may support the development of reagents that interfere with S100A4 function.
Malashkevich
Vladimir N
VN
Department of Biochemistry, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, New York 10461, USA.
Varney
Kristen M
KM
Garrett
Sarah C
SC
Wilder
Paul T
PT
Knight
David
D
Charpentier
Thomas H
TH
Ramagopal
Udupi A
UA
Almo
Steven C
SC
Weber
David J
DJ
Bresnick
Anne R
AR
D002118:Calcium / Q000378:metabolism*; D018360:Crystallography, X-Ray; D019281:Dimerization; D006801:Humans; D008958:Models, Molecular; D009132:Muscles / Q000737:chemistry / Q000378:metabolism; D024507:Nonmuscle Myosin Type IIA / Q000737:chemistry* / Q000378:metabolism*; D019906:Nuclear Magnetic Resonance, Biomolecular; D010446:Peptide Fragments / Q000737:chemistry* / Q000378:metabolism*; D011485:Protein Binding; D020836:Protein Structure, Quaternary; D000071999:S100 Calcium-Binding Protein A4; D009418:S100 Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D013816:Thermodynamics
D010446:Peptide Fragments; D000071999:S100 Calcium-Binding Protein A4; D009418:S100 Proteins; C077591:S100A4 protein, human; D024507:Nonmuscle Myosin Type IIA; D002118:Calcium