Supporting Information
© Copyright Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, 2008
© Copyright Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, 2008
Supporting Information
for
Globotriose-Functionalized Gold Nanoparticles as Multivalent Probes
for Shiga-like Toxin
Yuh-Yih Chien, Mi-Dan Jan, Avijit Kumar Adak, Hsiao-Chien Tzeng,
Yen-Ping Lin, Yu-Ju Chen, Ken-Tseng Wang, Chien-Tien Chen,
Chia-Chun Chen,* and Chun-Cheng Lin*
Contents
1. Figure S1. The UV-visible spectra of AuNPs S2
2. Figure S2. The X-ray structure of B-Slt and distance between two Pk
antigen binding sites S2
3. Figure S3. The linker length of the Pk antigen S3
4. Figure S4. Native polyacrylamide gel electrophoretic analysis of AuNP
extracted B-Slt S3
5. Calculation of Pk number on a particular size of AuNP S4
6. The IC50 of Pk –AuNPs to B-Slt S5
7. NMR spectra of compounds 1 and 10 S6 – S15
400 450 500 550 600 650 700
0.0
0.2
0.4
0.6
0.8
1.0
Abs
Wavelength (nm)
4-Pk-s-AuNP 13-Pk-s-AuNP 20-Pk-s-AuNP
Figure S1. The UV-visible spectra of AuNPs
14Asite1
site2
site3
17A17A
Between two site 127A
Between two site 229A
19A
14Asite1
site2
site3
17A17A
Between two site 127A
Between two site 229A
19A
Figure S2. The X-ray structure of B-Slt and distance between two Pk antigen binding sites.
S2
Figure S3. The linker length of the Pk antigen
Figure S4. Native polyacrylamide gel electrophoretic analysis of AuNP extracted B-Slt. Lane 1:
Alcohol dehydrogenase; Lane 2: Enolase; Lane 3: B-Slt; Lane 4: Marker
Calculation of Pk number on a particular size of AuNP
The number of gold atom composing a particular size of AuNP is directly related to its diameter
(eq. 1).
Gold atom number = 31.05* (diameter of AuNP)^3.0007 …………………………. (1)
S3
O
O O
HO
HOHOO
HOHO
OHO
OHO
NH
O
OH
OHOH
O
O O
HO
HOHOO
HOHO
OHO
OHO
NH
O
OH
OHOH
S
OO
O S
27.7A
13.3A
1 2 3 4
From eq 1, number of gold atom composing a particular size of AuNP can be calculated. For
example, total numbers of gold atoms for 2 and 10 nm size AuNPs are ~ 249 and ~ 31,097,
respectively. The number of gold atom of different size AuNP is given in Table 1.
Table 1.
Diameter (nm) Total atoms
4 1,989
13 68,339
20 248,776
Because the gold atom number composing a particular size of AuNP is known, the molecular weight
of this particular size AuNP can be calculated.
The amount of sugar on AuNP can be easily determined by anthrone-sulfuric acid assay. Briefly,
a standard curve was plotted for galactose using different concentrations ranging between 20 to 1 µg
in deionized water. A freshly prepared 0.5% anthrone (w/w) in 95% sulfuric acid was added to 1 mL
of sugar solutions. The mixture was carefully heated to 100 oC for 10 min and then allowed to cool.
The absorbance was recorded at 620 nm. Similarly, absorbances of glyconanoparticles (100 to 500
μg) were also recorded. From the standard curve, Pk ligand on nanoparticle was calculated and these
are given in Table 2. Combining the information of Tables 1 and 2, the number of Pk antigen on each
AuNP can be estimated.
Table 2.
Abbreviation Amount of sugar on 1 mg of Pk-AuNP
(µg/mg)
No. of sugar per nanoparticle
4-Pk-s-AuNP 90 60
13-Pk-s-AuNP 37.8 826
20-Pk-s-AuNP 21 1,498
4-Pk-l-AuNP 187 113
13-Pk-l-AuNP 67 1,298
20-Pk-l-AuNP 31.2 1,970
S4
The IC50 of Pk –AuNPs to B-Slt
The IC50 of Pk –AuNPs to B-Slt was estimated by using the Cheng-Prusoff equation1 (eq. 2).
IC50 = Ki * (1+ F/Kd) = Ki * (1+3.1E-8*1.16E8) = Ki *(1+3.596)………………(2)
Where the parameters are defined as: Kd is dissociation constant (1/Kd = Ka = 1.16E8 M-1), F is
concentration of the B-Slt (F = 3.1E-8).
Compound Ki (M)
Pk
4-s-Pk-AuNP 1.46 x 10-9
13-s-Pk-AuNP 8.64 x 10-11
20-s-Pk-AuNP 1.18 x 10-12
4-l-Pk-AuNP 1.95 x 10-11
13-l-Pk-AuNP
20-l-Pk-AuNP
6.84 x 10-13
2.54 x 10-13
Pk-s-dimer 3.68 x 10-5
Pk-l- dimer 6.38 x 10-6
1.14 x 10-4
IC50
5.11 x 10-4
1.65 x 10-4
2.86 x 10-5
6.54 x 10-9
3.87 x 10-10
5.29 x 10-12
8.74 x 10-11
3.06 x 10-12
1.14 x 10-12
Reference:
1. Cheng, Y.; Prusoff, W. H. Biochem. Pharmacol. 1973, 22, 3099-3108.
S5
10 9 8 7 6 5 4 3 2 1 ppm
N3O
OO
S6
2030405060708090100110120130140150160170180190 ppm
N3O
OO
S7
10 9 8 7 6 5 4 3 2 1 0 ppm
H2NO
OO
S8
2030405060708090100110120130140150160170180190 ppm
H2NO
OO
S9
10 9 8 7 6 5 4 3 2 1 ppm
OAcO
OAc
OAcO
OAc
OAc
O O
O OAcO
OAcO
AcO OAc
AcO
NH
OO
O
S10
2030405060708090100110120130140150160170180190 ppm
OAcO
OAc
OAcO
OAc
OAc
O O
O OAcO
OAcO
AcO OAc
AcO
NH
OO
O
S11
10 9 8 7 6 5 4 3 2 1 ppm
OAcO
OAc
OAcO
OAc
OAc
O O
O OAcO
OAcO
AcO OAc
AcO
NH
OO
O SAc
S12
210 200 190 180 170 160 150 140 130 120 110 100 90 80 70 60 50 40 30 20 10 ppm
OAcO
OAc
OAcO
OAc
OAc
O O
O OAcO
OAcO
AcO OAc
AcO
NH
OO
O SAc
S13
10 9 8 7 6 5 4 3 2 1 ppm
OHO
OH
OHO
OH
OH
O O
O OHO
OHO
HO OH
HO
NH
OO
O SH
S14
2030405060708090100110120130140150160170180190 ppm
OHO
OH
OHO
OH
OH
O O
O OHO
OHO
HO OH
HO
NH
OO
O SH
S15