The ATCUN domain as a probe of intermolecular interactions: application to calmodulin-peptide complexes.
The utility of a three-residue Cu2+ binding motif (ATCUN domain) for studying intermolecular interactions is demonstrated. By comparing a set of 1H-15N correlation spectra recorded on complexes of calmodulin (CaM) and peptides with the ATCUN tag in the presence and absence of Cu2+ the two possible canonical binding orientations of the peptide can be rapidly distinguished. The methodology is confirmed with studies of complexes of CaM and peptides from myosin light chain kinase and CaM kinase kinase, for which high-resolution structures are available, and then applied to a complex with CaM kinase I for which structural data has not been obtained. The orientation of the CaM kinase I and myosin light chain kinase peptides are shown to be identical. In the case of a complex of CaM with a peptide for which structural information is not available, the present methodology, in combination with 1H-15N residual dipolar couplings measured on CaM, and the database of existing CaM-peptide structures, allows a homology model to be built rapidly and with confidence.
Mal
Tapas K
TK
Division of Molecular and Structural Biology, Ontario Cancer Institute and Department of Medical Biophysics, University of Toronto, Toronto, Ontario, M5G 2M9, Canada.
Ikura
Mitsuhiko
M
Kay
Lewis E
LE
D000595:Amino Acid Sequence; D054729:Calcium-Calmodulin-Dependent Protein Kinase Type 1; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000737:chemistry* / Q000378:metabolism; D002147:Calmodulin / Q000737:chemistry* / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009219:Myosin-Light-Chain Kinase / Q000737:chemistry* / Q000378:metabolism; D019906:Nuclear Magnetic Resonance, Biomolecular / Q000379:methods; D011487:Protein Conformation; D017434:Protein Structure, Tertiary
D002147:Calmodulin; D054729:Calcium-Calmodulin-Dependent Protein Kinase Type 1; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D009219:Myosin-Light-Chain Kinase