Polymerization of calsequestrin. Implications for Ca2+ regulation.
Two distinct dimerization contacts in calsequestrin crystals suggested a mechanism for Ca(2+) regulation resulting from the occurrence of coupled Ca(2+) binding and protein polymerization. Ca(2+)-induced formation of one contact was proposed to lead to dimerization followed by Ca(2+)-induced formation of the second contact to bring about polymerization (). To test this mechanism, we compared canine cardiac calsequestrin and four truncation mutants with regard to their folding properties, structures, and Ca(2+)-induced polymerization. The wild-type calsequestrin and truncation mutants exhibited similar K(+)-induced folding and end-point structures as indicated by intrinsic fluorescence and circular dichroism, respectively, whereas the polymerization tendencies of the wild-type calsequestrin differed markedly from the polymerization tendencies of the truncation mutants. Static laser light scattering and 3,3'-dithiobis sulfosuccinimidyl-propionate cross-linking indicated that wild-type protein exhibited an initial Ca(2+)-induced dimerization, followed by additional oligomerization as the Ca(2+) concentration was raised or as the K(+) concentration was lowered. None of the truncation mutants exhibited clear stepwise oligomerization that depended on increasing Ca(2+) concentration. Comparison of the three-dimensional structure of rabbit skeletal calsequestrin with a homology model of canine cardiac calsequestrin from the point of view of our coupled Ca(2+) binding and polymerization mechanism leads to a possible explanation for the 2-fold reduced Ca(2+) binding capacity of cardiac calsequestrin despite very similar overall net negative charge for the two proteins.
Park
HaJeung
H
School of Molecular Biosciences, Department of Chemistry, Washington State University, Pullman, WA 99164, USA.
Wu
Si
S
Dunker
A Keith
AK
Kang
ChulHee
C
D000595:Amino Acid Sequence; D002118:Calcium / Q000378:metabolism*; D002155:Calsequestrin / Q000737:chemistry*; D002942:Circular Dichroism; D019281:Dimerization; D008027:Light; D008969:Molecular Sequence Data; D017510:Protein Folding; D012542:Scattering, Radiation; D013050:Spectrometry, Fluorescence
D002155:Calsequestrin; D002118:Calcium