Binding of Cu(II) to human alpha-synucleins: comparison of wild type and the point mutations associated with the familial Parkinson's disease.
The Cu(II)-alpha-synuclein interaction has been invoked as an important process in the pathogenesis of Parkinson's disease. Herein, we report binding constants and stoichiometry under near-physiological conditions for the binding of Cu(II) to human alpha-synuclein. Specifically, we compare the binding of Cu(II) to wild-type (WT) protein and two separate single mutation proteins that are associated with familial Parkinson's diseases: A30P and A53T. Cu(II) binds to all three alpha-synuclein proteins with a 1:1 stoichiometry. The Cu(II) binding constants, however, vary among the proteins studied. Cu(II) binding to WT and A53T at 37 degrees C is similar with a pH-dependent binding constant (K) of approximately 2.4 x 10(9) and approximately 4.8 x 10(9) M(-1) at pH 7.2 and 7.4, respectively. Cu(II) binding to A30P, however, exhibits two binding constants. The major binding site of A30P, characteristic of >90% of the bound Cu(II), has binding constants of 1.6 x 10(9) and 3.6 x 10(9) M(-1) at pH 7.2 and 7.4, respectively, slightly lower ( approximately 70%) than that characteristic of WT or A53T at the corresponding pH. The second less populated binding exhibited by A30P has a large binding constant, approximately 10(10) M(-1). Our size exclusion analysis ruled out the contribution of protofibrils to the strong Cu(II) binding. Previous studies indicated that A30P had a larger proportion of intermediate species (e.g., small oligomeric species, such as dimers and trimers) relative to WT and A53T. Thus, we propose that the high affinity site is attributed to the binding of Cu(II) to those small oligomeric species.
Hong
Lian
L
Department of Chemistry, Duke University, Durham, NC 27708, USA.
Simon
John D
JD
D000595:Amino Acid Sequence; D003300:Copper / Q000378:metabolism*; D006801:Humans; D008969:Molecular Sequence Data; D010300:Parkinson Disease / Q000235:genetics* / Q000378:metabolism*; D017354:Point Mutation; D011485:Protein Binding; D021241:Spectrometry, Mass, Electrospray Ionization; D051844:alpha-Synuclein / Q000235:genetics* / Q000378:metabolism*
C497604:SNCA protein, human; D051844:alpha-Synuclein; D003300:Copper