Intrinsically disordered PEP-19 confers unique dynamic properties to apo and calcium calmodulin.
PEP-19 (Purkinje cell protein 4) is an intrinsically disordered protein with an IQ calmodulin (CaM) binding motif. Expression of PEP-19 was recently shown to protect cells from apoptosis and cell death due to Ca(2+) overload. Our initial studies showed that PEP-19 causes novel and dramatic increases in the rates of association of Ca(2+) with and dissociation of Ca(2+) from the C-domain of CaM. The goal of this work was to study interactions between the C-domain of CaM (C-CaM) and PEP-19 by solution nuclear magnetic resonance (NMR) to identify mechanisms by which PEP-19 regulates binding of Ca(2+) to CaM. Our results show that PEP-19 causes a greater structural change in apo C-CaM than in Ca(2+)-C-CaM, and that the first Ca(2+) binds preferentially to site IV in the presence of PEP-19 with exchange characteristics that are consistent with a decrease in Ca(2+) binding cooperativity. Relatively weak binding of PEP-19 has distinct effects on chemical and conformational exchange on the microsecond to millisecond time scale. In apo C-CaM, PEP-19 binding causes a redistribution of residues that experience conformational exchange, leading to an increase in the number of residues around Ca(2+) binding site IV that undergo conformational exchange on the microsecond to millisecond time scale. This appears to be caused by an allosteric effect because these residues are not localized to the PEP-19 binding site. In contrast, PEP-19 increases the number of residues that exhibit conformational exchange in Ca(2+)-C-CaM. These residues are primarily localized to the PEP-19 binding site but also include Asp93 in site III. These results provide working models for the role of protein dynamics in the regulation of binding of Ca(2+) to CaM by PEP-19.
Wang
Xu
X
Department of Biochemistry and Molecular Biology and Structural Biology Center, University of Texas-Houston Medical School, 6431 Fannin Street, Houston, TX 77030, USA.
Kleerekoper
Quinn K
QK
Xiong
Liang-wen
LW
Putkey
John A
JA
D001059:Apoproteins / Q000737:chemistry / Q000378:metabolism*; D001665:Binding Sites; D002118:Calcium / Q000378:metabolism*; D002147:Calmodulin / Q000737:chemistry / Q000378:metabolism*; D007700:Kinetics; D008958:Models, Molecular; D009419:Nerve Tissue Proteins / Q000737:chemistry* / Q000378:metabolism*; D011485:Protein Binding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D015203:Reproducibility of Results
D001059:Apoproteins; D002147:Calmodulin; D009419:Nerve Tissue Proteins; D002118:Calcium