|
|
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result sql = Role of conformational change in the C-terminus of beta2-microglobulin in dialysis-related amyloidosis.
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char = n;code = 110
char = f;code = 102
char = o;code = 111
char = r;code = 114
char = m;code = 109
char = a;code = 97
char = t;code = 116
char = i;code = 105
char = o;code = 111
char = n;code = 110
char = a;code = 97
char = l;code = 108
char = ;code = 32
char = c;code = 99
char = h;code = 104
char = a;code = 97
char = n;code = 110
char = g;code = 103
char = e;code = 101
char = ;code = 32
char = a;code = 97
char = n;code = 110
char = d;code = 100
char = ;code = 32
char = a;code = 97
char = m;code = 109
char = y;code = 121
char = l;code = 108
char = o;code = 111
char = i;code = 105
char = d;code = 100
char = o;code = 111
char = g;code = 103
char = e;code = 101
char = n;code = 110
char = i;code = 105
char = c;code = 99
char = i;code = 105
char = t;code = 116
char = y;code = 121
char = ,;code = 44
char = ;code = 32
char = w;code = 119
char = e;code = 101
char = ;code = 32
char = m;code = 109
char = e;code = 101
char = a;code = 97
char = s;code = 115
char = u;code = 117
char = r;code = 114
char = e;code = 101
char = d;code = 100
char = ;code = 32
char = c;code = 99
char = i;code = 105
char = r;code = 114
char = c;code = 99
char = u;code = 117
char = l;code = 108
char = a;code = 97
char = r;code = 114
char = ;code = 32
char = d;code = 100
char = i;code = 105
char = c;code = 99
char = h;code = 104
char = r;code = 114
char = o;code = 111
char = i;code = 105
char = s;code = 115
char = m;code = 109
char = ;code = 32
char = s;code = 115
char = p;code = 112
char = e;code = 101
char = c;code = 99
char = t;code = 116
char = r;code = 114
char = a;code = 97
char = ,;code = 44
char = ;code = 32
char = t;code = 116
char = h;code = 104
char = e;code = 101
char = ;code = 32
char = f;code = 102
char = l;code = 108
char = u;code = 117
char = o;code = 111
char = r;code = 114
char = e;code = 101
char = s;code = 115
char = c;code = 99
char = e;code = 101
char = n;code = 110
char = c;code = 99
char = e;code = 101
char = ;code = 32
char = i;code = 105
char = n;code = 110
char = t;code = 116
char = e;code = 101
char = n;code = 110
char = s;code = 115
char = i;code = 105
char = t;code = 116
char = y;code = 121
char = ;code = 32
char = o;code = 111
char = f;code = 102
char = ;code = 32
char = t;code = 116
char = r;code = 114
char = y;code = 121
char = p;code = 112
char = t;code = 116
char = o;code = 111
char = p;code = 112
char = h;code = 104
char = a;code = 97
char = n;code = 110
char = ;code = 32
char = a;code = 97
char = n;code = 110
char = d;code = 100
char = ;code = 32
char = t;code = 116
char = h;code = 104
char = i;code = 105
char = o;code = 111
char = f;code = 102
char = l;code = 108
char = a;code = 97
char = v;code = 118
char = i;code = 105
char = n;code = 110
char = -;code = 45
char = T;code = 84
char = ;code = 32
char = (;code = 40
char = T;code = 84
char = h;code = 104
char = T;code = 84
char = );code = 41
char = ;code = 32
char = o;code = 111
char = f;code = 102
char = ;code = 32
char = t;code = 116
char = h;code = 104
char = e;code = 101
char = ;code = 32
char = r;code = 114
char = e;code = 101
char = c;code = 99
char = o;code = 111
char = m;code = 109
char = b;code = 98
char = i;code = 105
char = n;code = 110
char = a;code = 97
char = n;code = 110
char = t;code = 116
char = ;code = 32
char = b;code = 98
char = e;code = 101
char = t;code = 116
char = a;code = 97
char = (;code = 40
char = 2;code = 50
char = );code = 41
char = m;code = 109
char = .;code = 46
char = ;code = 32
char = T;code = 84
char = o;code = 111
char = ;code = 32
char = a;code = 97
char = n;code = 110
char = a;code = 97
char = l;code = 108
char = y;code = 121
char = s;code = 115
char = e;code = 101
char = ;code = 32
char = m;code = 109
char = o;code = 111
char = r;code = 114
char = p;code = 112
char = h;code = 104
char = o;code = 111
char = l;code = 108
char = o;code = 111
char = g;code = 103
char = i;code = 105
char = c;code = 99
char = a;code = 97
char = l;code = 108
char = ;code = 32
char = c;code = 99
char = h;code = 104
char = a;code = 97
char = n;code = 110
char = g;code = 103
char = e;code = 101
char = ;code = 32
char = o;code = 111
char = f;code = 102
char = ;code = 32
char = b;code = 98
char = e;code = 101
char = t;code = 116
char = a;code = 97
char = (;code = 40
char = 2;code = 50
char = );code = 41
char = m;code = 109
char = ,;code = 44
char = ;code = 32
char = w;code = 119
char = e;code = 101
char = ;code = 32
char = p;code = 112
char = e;code = 101
char = r;code = 114
char = f;code = 102
char = o;code = 111
char = r;code = 114
char = m;code = 109
char = e;code = 101
char = d;code = 100
char = ;code = 32
char = e;code = 101
char = l;code = 108
char = e;code = 101
char = c;code = 99
char = t;code = 116
char = r;code = 114
char = o;code = 111
char = n;code = 110
char = ;code = 32
char = m;code = 109
char = i;code = 105
char = c;code = 99
char = r;code = 114
char = o;code = 111
char = s;code = 115
char = c;code = 99
char = o;code = 111
char = p;code = 112
char = y;code = 121
char = ;code = 32
char = (;code = 40
char = E;code = 69
char = M;code = 77
char = );code = 41
char = ;code = 32
char = o;code = 111
char = n;code = 110
char = ;code = 32
char = t;code = 116
char = h;code = 104
char = e;code = 101
char = ;code = 32
char = s;code = 115
char = a;code = 97
char = m;code = 109
char = p;code = 112
char = l;code = 108
char = e;code = 101
char = s;code = 115
char = ;code = 32
char = w;code = 119
char = i;code = 105
char = t;code = 116
char = h;code = 104
char = ;code = 32
char = e;code = 101
char = l;code = 108
char = e;code = 101
char = v;code = 118
char = a;code = 97
char = t;code = 116
char = e;code = 101
char = d;code = 100
char = ;code = 32
char = T;code = 84
char = h;code = 104
char = T;code = 84
char = ;code = 32
char = f;code = 102
char = l;code = 108
char = u;code = 117
char = o;code = 111
char = r;code = 114
char = e;code = 101
char = s;code = 115
char = c;code = 99
char = e;code = 101
char = n;code = 110
char = c;code = 99
char = e;code = 101
char = ;code = 32
char = i;code = 105
char = n;code = 110
char = t;code = 116
char = e;code = 101
char = n;code = 110
char = s;code = 115
char = i;code = 105
char = t;code = 116
char = y;code = 121
char = .;code = 46
char = ;code = 32
char = W;code = 87
char = e;code = 101
char = ;code = 32
char = u;code = 117
char = s;code = 115
char = e;code = 101
char = d;code = 100
char = ;code = 32
char = u;code = 117
char = l;code = 108
char = t;code = 116
char = r;code = 114
char = a;code = 97
char = s;code = 115
char = o;code = 111
char = n;code = 110
char = i;code = 105
char = c;code = 99
char = a;code = 97
char = t;code = 116
char = i;code = 105
char = o;code = 111
char = n;code = 110
char = ;code = 32
char = t;code = 116
char = o;code = 111
char = ;code = 32
char = e;code = 101
char = n;code = 110
char = h;code = 104
char = a;code = 97
char = n;code = 110
char = c;code = 99
char = e;code = 101
char = ;code = 32
char = b;code = 98
char = e;code = 101
char = t;code = 116
char = a;code = 97
char = (;code = 40
char = 2;code = 50
char = );code = 41
char = m;code = 109
char = ;code = 32
char = d;code = 100
char = e;code = 101
char = s;code = 115
char = t;code = 116
char = a;code = 97
char = b;code = 98
char = i;code = 105
char = l;code = 108
char = i;code = 105
char = z;code = 122
char = a;code = 97
char = t;code = 116
char = i;code = 105
char = o;code = 111
char = n;code = 110
char = ;code = 32
char = o;code = 111
char = f;code = 102
char = ;code = 32
char = t;code = 116
char = h;code = 104
char = e;code = 101
char = ;code = 32
char = p;code = 112
char = r;code = 114
char = o;code = 111
char = t;code = 116
char = e;code = 101
char = i;code = 105
char = n;code = 110
char = .;code = 46
char = <;code = 60
char = b;code = 98
char = r;code = 114
char = /;code = 47
char = >;code = 62
char = <;code = 60
char = b;code = 98
char = r;code = 114
char = /;code = 47
char = >;code = 62
char = <;code = 60
char = b;code = 98
char = >;code = 62
char = R;code = 82
char = E;code = 69
char = S;code = 83
char = U;code = 85
char = L;code = 76
char = T;code = 84
char = S;code = 83
char = ;code = 32
char = :;code = 58
char = ;code = 32
char = <;code = 60
char = /;code = 47
char = b;code = 98
char = >;code = 62
char = B;code = 66
char = e;code = 101
char = t;code = 116
char = a;code = 97
char = (;code = 40
char = 2;code = 50
char = );code = 41
char = M;code = 77
char = ;code = 32
char = T;code = 84
char = r;code = 114
char = p;code = 112
char = 9;code = 57
char = 5;code = 53
char = L;code = 76
char = e;code = 101
char = u;code = 117
char = ;code = 32
char = a;code = 97
char = n;code = 110
char = d;code = 100
char = ;code = 32
char = A;code = 65
char = r;code = 114
char = g;code = 103
char = 9;code = 57
char = 7;code = 55
char = A;code = 65
char = l;code = 108
char = a;code = 97
char = ;code = 32
char = s;code = 115
char = h;code = 104
char = o;code = 111
char = w;code = 119
char = e;code = 101
char = d;code = 100
char = ;code = 32
char = c;code = 99
char = o;code = 111
char = n;code = 110
char = f;code = 102
char = o;code = 111
char = r;code = 114
char = m;code = 109
char = a;code = 97
char = t;code = 116
char = i;code = 105
char = o;code = 111
char = n;code = 110
char = a;code = 97
char = l;code = 108
char = ;code = 32
char = c;code = 99
char = h;code = 104
char = a;code = 97
char = n;code = 110
char = g;code = 103
char = e;code = 101
char = s;code = 115
char = ;code = 32
char = a;code = 97
char = n;code = 110
char = d;code = 100
char = ;code = 32
char = i;code = 105
char = n;code = 110
char = c;code = 99
char = r;code = 114
char = e;code = 101
char = a;code = 97
char = s;code = 115
char = e;code = 101
char = d;code = 100
char = ;code = 32
char = t;code = 116
char = h;code = 104
char = e;code = 101
char = i;code = 105
char = r;code = 114
char = ;code = 32
char = a;code = 97
char = m;code = 109
char = y;code = 121
char = l;code = 108
char = o;code = 111
char = i;code = 105
char = d;code = 100
char = g;code = 103
char = e;code = 101
char = n;code = 110
char = i;code = 105
char = c;code = 99
char = i;code = 105
char = t;code = 116
char = y;code = 121
char = ;code = 32
char = c;code = 99
char = o;code = 111
char = m;code = 109
char = p;code = 112
char = a;code = 97
char = r;code = 114
char = e;code = 101
char = d;code = 100
char = ;code = 32
char = w;code = 119
char = i;code = 105
char = t;code = 116
char = h;code = 104
char = ;code = 32
char = b;code = 98
char = e;code = 101
char = t;code = 116
char = a;code = 97
char = (;code = 40
char = 2;code = 50
char = );code = 41
char = m;code = 109
char = ;code = 32
char = w;code = 119
char = i;code = 105
char = l;code = 108
char = d;code = 100
char = -;code = 45
char = t;code = 116
char = y;code = 121
char = p;code = 112
char = e;code = 101
char = ;code = 32
char = (;code = 40
char = W;code = 87
char = T;code = 84
char = );code = 41
char = .;code = 46
char = ;code = 32
char = W;code = 87
char = i;code = 105
char = t;code = 116
char = h;code = 104
char = ;code = 32
char = u;code = 117
char = l;code = 108
char = t;code = 116
char = r;code = 114
char = a;code = 97
char = s;code = 115
char = o;code = 111
char = n;code = 110
char = i;code = 105
char = c;code = 99
char = a;code = 97
char = t;code = 116
char = i;code = 105
char = o;code = 111
char = n;code = 110
char = ,;code = 44
char = ;code = 32
char = b;code = 98
char = e;code = 101
char = t;code = 116
char = a;code = 97
char = (;code = 40
char = 2;code = 50
char = );code = 41
char = m;code = 109
char = ;code = 32
char = T;code = 84
char = r;code = 114
char = p;code = 112
char = 9;code = 57
char = 5;code = 53
char = L;code = 76
char = e;code = 101
char = u;code = 117
char = ;code = 32
char = a;code = 97
char = n;code = 110
char = d;code = 100
char = ;code = 32
char = A;code = 65
char = r;code = 114
char = g;code = 103
char = 9;code = 57
char = 7;code = 55
char = A;code = 65
char = l;code = 108
char = a;code = 97
char = ;code = 32
char = g;code = 103
char = e;code = 101
char = n;code = 110
char = e;code = 101
char = r;code = 114
char = a;code = 97
char = t;code = 116
char = e;code = 101
char = d;code = 100
char = ;code = 32
char = m;code = 109
char = o;code = 111
char = r;code = 114
char = e;code = 101
char = ;code = 32
char = a;code = 97
char = m;code = 109
char = y;code = 121
char = l;code = 108
char = o;code = 111
char = i;code = 105
char = d;code = 100
char = ;code = 32
char = f;code = 102
char = i;code = 105
char = b;code = 98
char = r;code = 114
char = i;code = 105
char = l;code = 108
char = s;code = 115
char = ;code = 32
char = t;code = 116
char = h;code = 104
char = a;code = 97
char = n;code = 110
char = ;code = 32
char = d;code = 100
char = i;code = 105
char = d;code = 100
char = ;code = 32
char = b;code = 98
char = e;code = 101
char = t;code = 116
char = a;code = 97
char = (;code = 40
char = 2;code = 50
char = );code = 41
char = m;code = 109
char = ;code = 32
char = W;code = 87
char = T;code = 84
char = ;code = 32
char = e;code = 101
char = v;code = 118
char = e;code = 101
char = n;code = 110
char = ;code = 32
char = i;code = 105
char = n;code = 110
char = ;code = 32
char = p;code = 112
char = h;code = 104
char = y;code = 121
char = s;code = 115
char = i;code = 105
char = o;code = 111
char = l;code = 108
char = o;code = 111
char = g;code = 103
char = i;code = 105
char = c;code = 99
char = a;code = 97
char = l;code = 108
char = ;code = 32
char = s;code = 115
char = o;code = 111
char = l;code = 108
char = u;code = 117
char = t;code = 116
char = i;code = 105
char = o;code = 111
char = n;code = 110
char = .;code = 46
char = ;code = 32
char = E;code = 69
char = M;code = 77
char = ;code = 32
char = s;code = 115
char = h;code = 104
char = o;code = 111
char = w;code = 119
char = e;code = 101
char = d;code = 100
char = ;code = 32
char = t;code = 116
char = h;code = 104
char = a;code = 97
char = t;code = 116
char = ;code = 32
char = b;code = 98
char = e;code = 101
char = t;code = 116
char = a;code = 97
char = (;code = 40
char = 2;code = 50
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char = m;code = 109
char = ;code = 32
char = f;code = 102
char = o;code = 111
char = r;code = 114
char = m;code = 109
char = e;code = 101
char = d;code = 100
char = ;code = 32
char = a;code = 97
char = m;code = 109
char = o;code = 111
char = r;code = 114
char = p;code = 112
char = h;code = 104
char = o;code = 111
char = u;code = 117
char = s;code = 115
char = ;code = 32
char = d;code = 100
char = e;code = 101
char = b;code = 98
char = r;code = 114
char = i;code = 105
char = s;code = 115
char = ;code = 32
char = c;code = 99
char = o;code = 111
char = n;code = 110
char = t;code = 116
char = a;code = 97
char = i;code = 105
char = n;code = 110
char = i;code = 105
char = n;code = 110
char = g;code = 103
char = ;code = 32
char = t;code = 116
char = y;code = 121
char = p;code = 112
char = i;code = 105
char = c;code = 99
char = a;code = 97
char = l;code = 108
char = ;code = 32
char = a;code = 97
char = m;code = 109
char = y;code = 121
char = l;code = 108
char = o;code = 111
char = i;code = 105
char = d;code = 100
char = ;code = 32
char = f;code = 102
char = i;code = 105
char = b;code = 98
char = r;code = 114
char = i;code = 105
char = l;code = 108
char = s;code = 115
char = ;code = 32
char = a;code = 97
char = t;code = 116
char = ;code = 32
char = 2;code = 50
char = 4;code = 52
char = ;code = 32
char = h;code = 104
char = o;code = 111
char = u;code = 117
char = r;code = 114
char = s;code = 115
char = ,;code = 44
char = ;code = 32
char = w;code = 119
char = h;code = 104
char = e;code = 101
char = n;code = 110
char = ;code = 32
char = T;code = 84
char = h;code = 104
char = T;code = 84
char = ;code = 32
char = f;code = 102
char = l;code = 108
char = u;code = 117
char = o;code = 111
char = r;code = 114
char = e;code = 101
char = s;code = 115
char = c;code = 99
char = e;code = 101
char = n;code = 110
char = c;code = 99
char = e;code = 101
char = ;code = 32
char = i;code = 105
char = n;code = 110
char = t;code = 116
char = e;code = 101
char = n;code = 110
char = s;code = 115
char = i;code = 105
char = t;code = 116
char = y;code = 121
char = ;code = 32
char = w;code = 119
char = a;code = 97
char = s;code = 115
char = ;code = 32
char = t;code = 116
char = h;code = 104
char = r;code = 114
char = e;code = 101
char = e;code = 101
char = -;code = 45
char = f;code = 102
char = o;code = 111
char = l;code = 108
char = d;code = 100
char = ;code = 32
char = l;code = 108
char = o;code = 111
char = w;code = 119
char = e;code = 101
char = r;code = 114
char = ;code = 32
char = t;code = 116
char = h;code = 104
char = a;code = 97
char = n;code = 110
char = ;code = 32
char = t;code = 116
char = h;code = 104
char = a;code = 97
char = t;code = 116
char = ;code = 32
char = a;code = 97
char = t;code = 116
char = ;code = 32
char = s;code = 115
char = i;code = 105
char = x;code = 120
char = ;code = 32
char = h;code = 104
char = o;code = 111
char = u;code = 117
char = r;code = 114
char = s;code = 115
char = .;code = 46
char = <;code = 60
char = b;code = 98
char = r;code = 114
char = /;code = 47
char = >;code = 62
char = <;code = 60
char = b;code = 98
char = r;code = 114
char = /;code = 47
char = >;code = 62
char = <;code = 60
char = b;code = 98
char = >;code = 62
char = C;code = 67
char = O;code = 79
char = N;code = 78
char = C;code = 67
char = L;code = 76
char = U;code = 85
char = S;code = 83
char = I;code = 73
char = O;code = 79
char = N;code = 78
char = S;code = 83
char = ;code = 32
char = :;code = 58
char = ;code = 32
char = <;code = 60
char = /;code = 47
char = b;code = 98
char = >;code = 62
char = C;code = 67
char = o;code = 111
char = n;code = 110
char = f;code = 102
char = o;code = 111
char = r;code = 114
char = m;code = 109
char = a;code = 97
char = t;code = 116
char = i;code = 105
char = o;code = 111
char = n;code = 110
char = a;code = 97
char = l;code = 108
char = ;code = 32
char = c;code = 99
char = h;code = 104
char = a;code = 97
char = n;code = 110
char = g;code = 103
char = e;code = 101
char = s;code = 115
char = ;code = 32
char = i;code = 105
char = n;code = 110
char = ;code = 32
char = t;code = 116
char = h;code = 104
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char = f;code = 102
char = u;code = 117
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char = f;code = 102
char = o;code = 111
char = r;code = 114
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char = a;code = 97
char = l;code = 108
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char = o;code = 111
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char = ;code = 32
char = b;code = 98
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char = t;code = 116
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char = 2;code = 50
char = );code = 41
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char = ;code = 32
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char = m;code = 109
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char = d;code = 100
char = o;code = 111
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result sql = BACKGROUND : Beta(2)-microglobulin (beta(2)m) has been identified as the precursor protein of dialysis-related amyloidosis (DRA), which is a serious complication for haemodialysis (HD) patients. However, mechanisms underlying beta(2)m amyloid fibril formation remains to be elucidated. We previously demonstrated, in amyloid deposits from HD patients, a conformational isoform of beta(2)m with an unfolded C-terminus. However, no direct experiments have previously been performed to address whether unfolded beta(2)m in the C-terminus may be prone to form amyloid fibrils.
METHODS : To evaluate roles of C-terminal amino acids in beta(2)m-induced amyloid formation, we generated six types of recombinant beta(2)m with amino acid substitutions in the C-terminal region. To investigate their conformational change and amyloidogenicity, we measured circular dichroism spectra, the fluorescence intensity of tryptophan and thioflavin-T (ThT) of the recombinant beta(2)m. To analyse morphological change of beta(2)m, we performed electron microscopy (EM) on the samples with elevated ThT fluorescence intensity. We used ultrasonication to enhance beta(2)m destabilization of the protein.
RESULTS : Beta(2)M Trp95Leu and Arg97Ala showed conformational changes and increased their amyloidgenicity compared with beta(2)m wild-type (WT). With ultrasonication, beta(2)m Trp95Leu and Arg97Ala generated more amyloid fibrils than did beta(2)m WT even in physiological solution. EM showed that beta(2)m formed amorphous debris containing typical amyloid fibrils at 24 hours, when ThT fluorescence intensity was three-fold lower than that at six hours.
CONCLUSIONS : Conformational changes in the C-terminus of beta(2)m may play an important role in DRA and that ultrasonication is useful for analysis of beta(2)m amyloidogenesis.
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char = n;code = 110
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char = r;code = 114
char = m;code = 109
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char = t;code = 116
char = i;code = 105
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char = c;code = 99
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char = n;code = 110
char = g;code = 103
char = e;code = 101
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char = i;code = 105
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char = h;code = 104
char = e;code = 101
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char = g;code = 103
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char = d;code = 100
char = o;code = 111
char = s;code = 115
char = i;code = 105
char = s;code = 115
char = .;code = 46
result sql = Role of conformational change in the C-terminus of beta2-microglobulin in dialysis-related amyloidosis.
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char = a;code = 97
char = m;code = 109
char = i;code = 105
char = n;code = 110
char = o;code = 111
char = ;code = 32
char = a;code = 97
char = c;code = 99
char = i;code = 105
char = d;code = 100
char = s;code = 115
char = ;code = 32
char = i;code = 105
char = n;code = 110
char = ;code = 32
char = b;code = 98
char = e;code = 101
char = t;code = 116
char = a;code = 97
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char = 2;code = 50
char = );code = 41
char = m;code = 109
char = -;code = 45
char = i;code = 105
char = n;code = 110
char = d;code = 100
char = u;code = 117
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char = e;code = 101
char = d;code = 100
char = ;code = 32
char = a;code = 97
char = m;code = 109
char = y;code = 121
char = l;code = 108
char = o;code = 111
char = i;code = 105
char = d;code = 100
char = ;code = 32
char = f;code = 102
char = o;code = 111
char = r;code = 114
char = m;code = 109
char = a;code = 97
char = t;code = 116
char = i;code = 105
char = o;code = 111
char = n;code = 110
char = ,;code = 44
char = ;code = 32
char = w;code = 119
char = e;code = 101
char = ;code = 32
char = g;code = 103
char = e;code = 101
char = n;code = 110
char = e;code = 101
char = r;code = 114
char = a;code = 97
char = t;code = 116
char = e;code = 101
char = d;code = 100
char = ;code = 32
char = s;code = 115
char = i;code = 105
char = x;code = 120
char = ;code = 32
char = t;code = 116
char = y;code = 121
char = p;code = 112
char = e;code = 101
char = s;code = 115
char = ;code = 32
char = o;code = 111
char = f;code = 102
char = ;code = 32
char = r;code = 114
char = e;code = 101
char = c;code = 99
char = o;code = 111
char = m;code = 109
char = b;code = 98
char = i;code = 105
char = n;code = 110
char = a;code = 97
char = n;code = 110
char = t;code = 116
char = ;code = 32
char = b;code = 98
char = e;code = 101
char = t;code = 116
char = a;code = 97
char = (;code = 40
char = 2;code = 50
char = );code = 41
char = m;code = 109
char = ;code = 32
char = w;code = 119
char = i;code = 105
char = t;code = 116
char = h;code = 104
char = ;code = 32
char = a;code = 97
char = m;code = 109
char = i;code = 105
char = n;code = 110
char = o;code = 111
char = ;code = 32
char = a;code = 97
char = c;code = 99
char = i;code = 105
char = d;code = 100
char = ;code = 32
char = s;code = 115
char = u;code = 117
char = b;code = 98
char = s;code = 115
char = t;code = 116
char = i;code = 105
char = t;code = 116
char = u;code = 117
char = t;code = 116
char = i;code = 105
char = o;code = 111
char = n;code = 110
char = s;code = 115
char = ;code = 32
char = i;code = 105
char = n;code = 110
char = ;code = 32
char = t;code = 116
char = h;code = 104
char = e;code = 101
char = ;code = 32
char = C;code = 67
char = -;code = 45
char = t;code = 116
char = e;code = 101
char = r;code = 114
char = m;code = 109
char = i;code = 105
char = n;code = 110
char = a;code = 97
char = l;code = 108
char = ;code = 32
char = r;code = 114
char = e;code = 101
char = g;code = 103
char = i;code = 105
char = o;code = 111
char = n;code = 110
char = .;code = 46
char = ;code = 32
char = T;code = 84
char = o;code = 111
char = ;code = 32
char = i;code = 105
char = n;code = 110
char = v;code = 118
char = e;code = 101
char = s;code = 115
char = t;code = 116
char = i;code = 105
char = g;code = 103
char = a;code = 97
char = t;code = 116
char = e;code = 101
char = ;code = 32
char = t;code = 116
char = h;code = 104
char = e;code = 101
char = i;code = 105
char = r;code = 114
char = ;code = 32
char = c;code = 99
char = o;code = 111
char = n;code = 110
char = f;code = 102
char = o;code = 111
char = r;code = 114
char = m;code = 109
char = a;code = 97
char = t;code = 116
char = i;code = 105
char = o;code = 111
char = n;code = 110
char = a;code = 97
char = l;code = 108
char = ;code = 32
char = c;code = 99
char = h;code = 104
char = a;code = 97
char = n;code = 110
char = g;code = 103
char = e;code = 101
char = ;code = 32
char = a;code = 97
char = n;code = 110
char = d;code = 100
char = ;code = 32
char = a;code = 97
char = m;code = 109
char = y;code = 121
char = l;code = 108
char = o;code = 111
char = i;code = 105
char = d;code = 100
char = o;code = 111
char = g;code = 103
char = e;code = 101
char = n;code = 110
char = i;code = 105
char = c;code = 99
char = i;code = 105
char = t;code = 116
char = y;code = 121
char = ,;code = 44
char = ;code = 32
char = w;code = 119
char = e;code = 101
char = ;code = 32
char = m;code = 109
char = e;code = 101
char = a;code = 97
char = s;code = 115
char = u;code = 117
char = r;code = 114
char = e;code = 101
char = d;code = 100
char = ;code = 32
char = c;code = 99
char = i;code = 105
char = r;code = 114
char = c;code = 99
char = u;code = 117
char = l;code = 108
char = a;code = 97
char = r;code = 114
char = ;code = 32
char = d;code = 100
char = i;code = 105
char = c;code = 99
char = h;code = 104
char = r;code = 114
char = o;code = 111
char = i;code = 105
char = s;code = 115
char = m;code = 109
char = ;code = 32
char = s;code = 115
char = p;code = 112
char = e;code = 101
char = c;code = 99
char = t;code = 116
char = r;code = 114
char = a;code = 97
char = ,;code = 44
char = ;code = 32
char = t;code = 116
char = h;code = 104
char = e;code = 101
char = ;code = 32
char = f;code = 102
char = l;code = 108
char = u;code = 117
char = o;code = 111
char = r;code = 114
char = e;code = 101
char = s;code = 115
char = c;code = 99
char = e;code = 101
char = n;code = 110
char = c;code = 99
char = e;code = 101
char = ;code = 32
char = i;code = 105
char = n;code = 110
char = t;code = 116
char = e;code = 101
char = n;code = 110
char = s;code = 115
char = i;code = 105
char = t;code = 116
char = y;code = 121
char = ;code = 32
char = o;code = 111
char = f;code = 102
char = ;code = 32
char = t;code = 116
char = r;code = 114
char = y;code = 121
char = p;code = 112
char = t;code = 116
char = o;code = 111
char = p;code = 112
char = h;code = 104
char = a;code = 97
char = n;code = 110
char = ;code = 32
char = a;code = 97
char = n;code = 110
char = d;code = 100
char = ;code = 32
char = t;code = 116
char = h;code = 104
char = i;code = 105
char = o;code = 111
char = f;code = 102
char = l;code = 108
char = a;code = 97
char = v;code = 118
char = i;code = 105
char = n;code = 110
char = -;code = 45
char = T;code = 84
char = ;code = 32
char = (;code = 40
char = T;code = 84
char = h;code = 104
char = T;code = 84
char = );code = 41
char = ;code = 32
char = o;code = 111
char = f;code = 102
char = ;code = 32
char = t;code = 116
char = h;code = 104
char = e;code = 101
char = ;code = 32
char = r;code = 114
char = e;code = 101
char = c;code = 99
char = o;code = 111
char = m;code = 109
char = b;code = 98
char = i;code = 105
char = n;code = 110
char = a;code = 97
char = n;code = 110
char = t;code = 116
char = ;code = 32
char = b;code = 98
char = e;code = 101
char = t;code = 116
char = a;code = 97
char = (;code = 40
char = 2;code = 50
char = );code = 41
char = m;code = 109
char = .;code = 46
char = ;code = 32
char = T;code = 84
char = o;code = 111
char = ;code = 32
char = a;code = 97
char = n;code = 110
char = a;code = 97
char = l;code = 108
char = y;code = 121
char = s;code = 115
char = e;code = 101
char = ;code = 32
char = m;code = 109
char = o;code = 111
char = r;code = 114
char = p;code = 112
char = h;code = 104
char = o;code = 111
char = l;code = 108
char = o;code = 111
char = g;code = 103
char = i;code = 105
char = c;code = 99
char = a;code = 97
char = l;code = 108
char = ;code = 32
char = c;code = 99
char = h;code = 104
char = a;code = 97
char = n;code = 110
char = g;code = 103
char = e;code = 101
char = ;code = 32
char = o;code = 111
char = f;code = 102
char = ;code = 32
char = b;code = 98
char = e;code = 101
char = t;code = 116
char = a;code = 97
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char = 2;code = 50
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char = m;code = 109
char = ,;code = 44
char = ;code = 32
char = w;code = 119
char = e;code = 101
char = ;code = 32
char = p;code = 112
char = e;code = 101
char = r;code = 114
char = f;code = 102
char = o;code = 111
char = r;code = 114
char = m;code = 109
char = e;code = 101
char = d;code = 100
char = ;code = 32
char = e;code = 101
char = l;code = 108
char = e;code = 101
char = c;code = 99
char = t;code = 116
char = r;code = 114
char = o;code = 111
char = n;code = 110
char = ;code = 32
char = m;code = 109
char = i;code = 105
char = c;code = 99
char = r;code = 114
char = o;code = 111
char = s;code = 115
char = c;code = 99
char = o;code = 111
char = p;code = 112
char = y;code = 121
char = ;code = 32
char = (;code = 40
char = E;code = 69
char = M;code = 77
char = );code = 41
char = ;code = 32
char = o;code = 111
char = n;code = 110
char = ;code = 32
char = t;code = 116
char = h;code = 104
char = e;code = 101
char = ;code = 32
char = s;code = 115
char = a;code = 97
char = m;code = 109
char = p;code = 112
char = l;code = 108
char = e;code = 101
char = s;code = 115
char = ;code = 32
char = w;code = 119
char = i;code = 105
char = t;code = 116
char = h;code = 104
char = ;code = 32
char = e;code = 101
char = l;code = 108
char = e;code = 101
char = v;code = 118
char = a;code = 97
char = t;code = 116
char = e;code = 101
char = d;code = 100
char = ;code = 32
char = T;code = 84
char = h;code = 104
char = T;code = 84
char = ;code = 32
char = f;code = 102
char = l;code = 108
char = u;code = 117
char = o;code = 111
char = r;code = 114
char = e;code = 101
char = s;code = 115
char = c;code = 99
char = e;code = 101
char = n;code = 110
char = c;code = 99
char = e;code = 101
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char = i;code = 105
char = n;code = 110
char = t;code = 116
char = e;code = 101
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char = s;code = 115
char = i;code = 105
char = t;code = 116
char = y;code = 121
char = .;code = 46
char = ;code = 32
char = W;code = 87
char = e;code = 101
char = ;code = 32
char = u;code = 117
char = s;code = 115
char = e;code = 101
char = d;code = 100
char = ;code = 32
char = u;code = 117
char = l;code = 108
char = t;code = 116
char = r;code = 114
char = a;code = 97
char = s;code = 115
char = o;code = 111
char = n;code = 110
char = i;code = 105
char = c;code = 99
char = a;code = 97
char = t;code = 116
char = i;code = 105
char = o;code = 111
char = n;code = 110
char = ;code = 32
char = t;code = 116
char = o;code = 111
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char = e;code = 101
char = n;code = 110
char = h;code = 104
char = a;code = 97
char = n;code = 110
char = c;code = 99
char = e;code = 101
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char = b;code = 98
char = e;code = 101
char = t;code = 116
char = a;code = 97
char = (;code = 40
char = 2;code = 50
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char = m;code = 109
char = ;code = 32
char = d;code = 100
char = e;code = 101
char = s;code = 115
char = t;code = 116
char = a;code = 97
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char = i;code = 105
char = l;code = 108
char = i;code = 105
char = z;code = 122
char = a;code = 97
char = t;code = 116
char = i;code = 105
char = o;code = 111
char = n;code = 110
char = ;code = 32
char = o;code = 111
char = f;code = 102
char = ;code = 32
char = t;code = 116
char = h;code = 104
char = e;code = 101
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char = p;code = 112
char = r;code = 114
char = o;code = 111
char = t;code = 116
char = e;code = 101
char = i;code = 105
char = n;code = 110
char = .;code = 46
char = <;code = 60
char = b;code = 98
char = r;code = 114
char = /;code = 47
char = >;code = 62
char = <;code = 60
char = b;code = 98
char = r;code = 114
char = /;code = 47
char = >;code = 62
char = <;code = 60
char = b;code = 98
char = >;code = 62
char = R;code = 82
char = E;code = 69
char = S;code = 83
char = U;code = 85
char = L;code = 76
char = T;code = 84
char = S;code = 83
char = ;code = 32
char = :;code = 58
char = ;code = 32
char = <;code = 60
char = /;code = 47
char = b;code = 98
char = >;code = 62
char = B;code = 66
char = e;code = 101
char = t;code = 116
char = a;code = 97
char = (;code = 40
char = 2;code = 50
char = );code = 41
char = M;code = 77
char = ;code = 32
char = T;code = 84
char = r;code = 114
char = p;code = 112
char = 9;code = 57
char = 5;code = 53
char = L;code = 76
char = e;code = 101
char = u;code = 117
char = ;code = 32
char = a;code = 97
char = n;code = 110
char = d;code = 100
char = ;code = 32
char = A;code = 65
char = r;code = 114
char = g;code = 103
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char = 7;code = 55
char = A;code = 65
char = l;code = 108
char = a;code = 97
char = ;code = 32
char = s;code = 115
char = h;code = 104
char = o;code = 111
char = w;code = 119
char = e;code = 101
char = d;code = 100
char = ;code = 32
char = c;code = 99
char = o;code = 111
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char = f;code = 102
char = o;code = 111
char = r;code = 114
char = m;code = 109
char = a;code = 97
char = t;code = 116
char = i;code = 105
char = o;code = 111
char = n;code = 110
char = a;code = 97
char = l;code = 108
char = ;code = 32
char = c;code = 99
char = h;code = 104
char = a;code = 97
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char = g;code = 103
char = e;code = 101
char = s;code = 115
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char = d;code = 100
char = ;code = 32
char = i;code = 105
char = n;code = 110
char = c;code = 99
char = r;code = 114
char = e;code = 101
char = a;code = 97
char = s;code = 115
char = e;code = 101
char = d;code = 100
char = ;code = 32
char = t;code = 116
char = h;code = 104
char = e;code = 101
char = i;code = 105
char = r;code = 114
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char = a;code = 97
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char = y;code = 121
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char = e;code = 101
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char = p;code = 112
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char = t;code = 116
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char = b;code = 98
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char = t;code = 116
char = a;code = 97
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char = m;code = 109
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char = w;code = 119
char = i;code = 105
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char = d;code = 100
char = -;code = 45
char = t;code = 116
char = y;code = 121
char = p;code = 112
char = e;code = 101
char = ;code = 32
char = (;code = 40
char = W;code = 87
char = T;code = 84
char = );code = 41
char = .;code = 46
char = ;code = 32
char = W;code = 87
char = i;code = 105
char = t;code = 116
char = h;code = 104
char = ;code = 32
char = u;code = 117
char = l;code = 108
char = t;code = 116
char = r;code = 114
char = a;code = 97
char = s;code = 115
char = o;code = 111
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char = i;code = 105
char = c;code = 99
char = a;code = 97
char = t;code = 116
char = i;code = 105
char = o;code = 111
char = n;code = 110
char = ,;code = 44
char = ;code = 32
char = b;code = 98
char = e;code = 101
char = t;code = 116
char = a;code = 97
char = (;code = 40
char = 2;code = 50
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char = m;code = 109
char = ;code = 32
char = T;code = 84
char = r;code = 114
char = p;code = 112
char = 9;code = 57
char = 5;code = 53
char = L;code = 76
char = e;code = 101
char = u;code = 117
char = ;code = 32
char = a;code = 97
char = n;code = 110
char = d;code = 100
char = ;code = 32
char = A;code = 65
char = r;code = 114
char = g;code = 103
char = 9;code = 57
char = 7;code = 55
char = A;code = 65
char = l;code = 108
char = a;code = 97
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char = g;code = 103
char = e;code = 101
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char = e;code = 101
char = r;code = 114
char = a;code = 97
char = t;code = 116
char = e;code = 101
char = d;code = 100
char = ;code = 32
char = m;code = 109
char = o;code = 111
char = r;code = 114
char = e;code = 101
char = ;code = 32
char = a;code = 97
char = m;code = 109
char = y;code = 121
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char = o;code = 111
char = i;code = 105
char = d;code = 100
char = ;code = 32
char = f;code = 102
char = i;code = 105
char = b;code = 98
char = r;code = 114
char = i;code = 105
char = l;code = 108
char = s;code = 115
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char = t;code = 116
char = h;code = 104
char = a;code = 97
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char = d;code = 100
char = i;code = 105
char = d;code = 100
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char = b;code = 98
char = e;code = 101
char = t;code = 116
char = a;code = 97
char = (;code = 40
char = 2;code = 50
char = );code = 41
char = m;code = 109
char = ;code = 32
char = W;code = 87
char = T;code = 84
char = ;code = 32
char = e;code = 101
char = v;code = 118
char = e;code = 101
char = n;code = 110
char = ;code = 32
char = i;code = 105
char = n;code = 110
char = ;code = 32
char = p;code = 112
char = h;code = 104
char = y;code = 121
char = s;code = 115
char = i;code = 105
char = o;code = 111
char = l;code = 108
char = o;code = 111
char = g;code = 103
char = i;code = 105
char = c;code = 99
char = a;code = 97
char = l;code = 108
char = ;code = 32
char = s;code = 115
char = o;code = 111
char = l;code = 108
char = u;code = 117
char = t;code = 116
char = i;code = 105
char = o;code = 111
char = n;code = 110
char = .;code = 46
char = ;code = 32
char = E;code = 69
char = M;code = 77
char = ;code = 32
char = s;code = 115
char = h;code = 104
char = o;code = 111
char = w;code = 119
char = e;code = 101
char = d;code = 100
char = ;code = 32
char = t;code = 116
char = h;code = 104
char = a;code = 97
char = t;code = 116
char = ;code = 32
char = b;code = 98
char = e;code = 101
char = t;code = 116
char = a;code = 97
char = (;code = 40
char = 2;code = 50
char = );code = 41
char = m;code = 109
char = ;code = 32
char = f;code = 102
char = o;code = 111
char = r;code = 114
char = m;code = 109
char = e;code = 101
char = d;code = 100
char = ;code = 32
char = a;code = 97
char = m;code = 109
char = o;code = 111
char = r;code = 114
char = p;code = 112
char = h;code = 104
char = o;code = 111
char = u;code = 117
char = s;code = 115
char = ;code = 32
char = d;code = 100
char = e;code = 101
char = b;code = 98
char = r;code = 114
char = i;code = 105
char = s;code = 115
char = ;code = 32
char = c;code = 99
char = o;code = 111
char = n;code = 110
char = t;code = 116
char = a;code = 97
char = i;code = 105
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result sql = BACKGROUND : Beta(2)-microglobulin (beta(2)m) has been identified as the precursor protein of dialysis-related amyloidosis (DRA), which is a serious complication for haemodialysis (HD) patients. However, mechanisms underlying beta(2)m amyloid fibril formation remains to be elucidated. We previously demonstrated, in amyloid deposits from HD patients, a conformational isoform of beta(2)m with an unfolded C-terminus. However, no direct experiments have previously been performed to address whether unfolded beta(2)m in the C-terminus may be prone to form amyloid fibrils.
METHODS : To evaluate roles of C-terminal amino acids in beta(2)m-induced amyloid formation, we generated six types of recombinant beta(2)m with amino acid substitutions in the C-terminal region. To investigate their conformational change and amyloidogenicity, we measured circular dichroism spectra, the fluorescence intensity of tryptophan and thioflavin-T (ThT) of the recombinant beta(2)m. To analyse morphological change of beta(2)m, we performed electron microscopy (EM) on the samples with elevated ThT fluorescence intensity. We used ultrasonication to enhance beta(2)m destabilization of the protein.
RESULTS : Beta(2)M Trp95Leu and Arg97Ala showed conformational changes and increased their amyloidgenicity compared with beta(2)m wild-type (WT). With ultrasonication, beta(2)m Trp95Leu and Arg97Ala generated more amyloid fibrils than did beta(2)m WT even in physiological solution. EM showed that beta(2)m formed amorphous debris containing typical amyloid fibrils at 24 hours, when ThT fluorescence intensity was three-fold lower than that at six hours.
CONCLUSIONS : Conformational changes in the C-terminus of beta(2)m may play an important role in DRA and that ultrasonication is useful for analysis of beta(2)m amyloidogenesis.
Publication: Ann. Clin. Biochem. 2008 Sep ;45(Pt 5)::489-95. :doi: 10.1258/acb.2008.008046.
Publication Date: 2008 Sep
Study Author(s): Kim, Jaemi;Motomiya, Yoshihiro;Ueda, Mitsuharu;Nakamura, Masaaki;Misumi, Yohei;Saito, Shiori;Ikemizu, Shinji;Misumi, Shogo;Ota, Kazutoshi;Shinriki, Satoru;Kai, Hirofumi;Ando, Yukio;
Institution: Department of Molecular Medicine, Graduate School of Medical Sciences, Kumamoto University, 1-1-1 Honjo, Kumamoto, Japan.
Shortcut link to this study:
Publication Date: 2008 Sep
Study Author(s): Kim, Jaemi;Motomiya, Yoshihiro;Ueda, Mitsuharu;Nakamura, Masaaki;Misumi, Yohei;Saito, Shiori;Ikemizu, Shinji;Misumi, Shogo;Ota, Kazutoshi;Shinriki, Satoru;Kai, Hirofumi;Ando, Yukio;
Institution: Department of Molecular Medicine, Graduate School of Medical Sciences, Kumamoto University, 1-1-1 Honjo, Kumamoto, Japan.
Shortcut link to this study:
BACKGROUND : Beta(2)-microglobulin (beta(2)m) has been identified as the precursor protein of dialysis-related amyloidosis (DRA), which is a serious complication for haemodialysis (HD) patients. However, mechanisms underlying beta(2)m amyloid fibril formation remains to be elucidated. We previously demonstrated, in amyloid deposits from HD patients, a conformational isoform of beta(2)m with an unfolded C-terminus. However, no direct experiments have previously been performed to address whether unfolded beta(2)m in the C-terminus may be prone to form amyloid fibrils.
METHODS : To evaluate roles of C-terminal amino acids in beta(2)m-induced amyloid formation, we generated six types of recombinant beta(2)m with amino acid substitutions in the C-terminal region. To investigate their conformational change and amyloidogenicity, we measured circular dichroism spectra, the fluorescence intensity of tryptophan and thioflavin-T (ThT) of the recombinant beta(2)m. To analyse morphological change of beta(2)m, we performed electron microscopy (EM) on the samples with elevated ThT fluorescence intensity. We used ultrasonication to enhance beta(2)m destabilization of the protein.
RESULTS : Beta(2)M Trp95Leu and Arg97Ala showed conformational changes and increased their amyloidgenicity compared with beta(2)m wild-type (WT). With ultrasonication, beta(2)m Trp95Leu and Arg97Ala generated more amyloid fibrils than did beta(2)m WT even in physiological solution. EM showed that beta(2)m formed amorphous debris containing typical amyloid fibrils at 24 hours, when ThT fluorescence intensity was three-fold lower than that at six hours.
CONCLUSIONS : Conformational changes in the C-terminus of beta(2)m may play an important role in DRA and that ultrasonication is useful for analysis of beta(2)m amyloidogenesis.
METHODS : To evaluate roles of C-terminal amino acids in beta(2)m-induced amyloid formation, we generated six types of recombinant beta(2)m with amino acid substitutions in the C-terminal region. To investigate their conformational change and amyloidogenicity, we measured circular dichroism spectra, the fluorescence intensity of tryptophan and thioflavin-T (ThT) of the recombinant beta(2)m. To analyse morphological change of beta(2)m, we performed electron microscopy (EM) on the samples with elevated ThT fluorescence intensity. We used ultrasonication to enhance beta(2)m destabilization of the protein.
RESULTS : Beta(2)M Trp95Leu and Arg97Ala showed conformational changes and increased their amyloidgenicity compared with beta(2)m wild-type (WT). With ultrasonication, beta(2)m Trp95Leu and Arg97Ala generated more amyloid fibrils than did beta(2)m WT even in physiological solution. EM showed that beta(2)m formed amorphous debris containing typical amyloid fibrils at 24 hours, when ThT fluorescence intensity was three-fold lower than that at six hours.
CONCLUSIONS : Conformational changes in the C-terminus of beta(2)m may play an important role in DRA and that ultrasonication is useful for analysis of beta(2)m amyloidogenesis.
PMID: 18753422
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