Interaction between the C-terminal region of human myelin basic protein and calmodulin: analysis of complex formation and solution structure.
BACKGROUND
The myelin sheath is a multilamellar membrane structure wrapped around the axon, enabling the saltatory conduction of nerve impulses in vertebrates. Myelin basic protein, one of the most abundant myelin-specific proteins, is an intrinsically disordered protein that has been shown to bind calmodulin. In this study, we focus on a 19-mer synthetic peptide from the predicted calmodulin-binding segment near the C-terminus of human myelin basic protein.


RESULTS
The interaction of native human myelin basic protein with calmodulin was confirmed by affinity chromatography. The binding of the myelin basic protein peptide to calmodulin was tested with isothermal titration calorimetry (ITC) in different temperatures, and Kd was observed to be in the low muM range, as previously observed for full-length myelin basic protein. Surface plasmon resonance showed that the peptide bound to calmodulin, and binding was accompanied by a conformational change; furthermore, gel filtration chromatography indicated a decrease in the hydrodynamic radius of calmodulin in the presence of the peptide. NMR spectroscopy was used to map the binding area to reside mainly within the hydrophobic pocket of the C-terminal lobe of calmodulin. The solution structure obtained by small-angle X-ray scattering indicates binding of the myelin basic protein peptide into the interlobal groove of calmodulin, while calmodulin remains in an extended conformation.


CONCLUSIONS
Taken together, our results give a detailed structural insight into the interaction of calmodulin with a C-terminal segment of a major myelin protein, the myelin basic protein. The used 19-mer peptide interacts mainly with the C-terminal lobe of calmodulin, and a conformational change accompanies binding, suggesting a novel mode of calmodulin-target protein interaction. Calmodulin does not collapse and wrap around the peptide tightly; instead, it remains in an extended conformation in the solution structure. The observed affinity can be physiologically relevant, given the high abundance of both binding partners in the nervous system.
Majava
Viivi
V
Department of Biochemistry, University of Oulu, Oulu, Finland. viivi.majava@oulu.fi
Petoukhov
Maxim V
MV
Hayashi
Nobuhiro
N
Pirilä
Päivi
P
Svergun
Dmitri I
DI
Kursula
Petri
P
D000595:Amino Acid Sequence; D001665:Binding Sites; D001921:Brain / Q000378:metabolism; D002147:Calmodulin / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D002846:Chromatography, Affinity; D002850:Chromatography, Gel; D002942:Circular Dichroism; D018360:Crystallography, X-Ray; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D008958:Models, Molecular; D008969:Molecular Sequence Data; D004676:Myelin Basic Protein; D009419:Nerve Tissue Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D019906:Nuclear Magnetic Resonance, Biomolecular; D011485:Protein Binding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D020349:Surface Plasmon Resonance; D014157:Transcription Factors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
D002147:Calmodulin; C499048:MBP protein, human; D004676:Myelin Basic Protein; D009419:Nerve Tissue Proteins; D014157:Transcription Factors