their mutated derivatives The unusual metal ion binding ability of … · 2016. 2. 17. · E-mail: [email protected] b Department of Chemistry, University of Wroclaw, F. Joliot-Curie 14,

Supporting Information for

"The unusual metal ion binding ability of histidyl tags and

their mutated derivatives"

Davide Brasili,a Joanna Watly, b Eyal Simonovsky, cd Remo Guerrini,a Nuno A. Barbosa, b

Robert Wieczorekb, Maurizio Remellia, Henryk Kozlowskib* and Yifat Millercd*

a Department of Chemical and Pharmaceutical Sciences, University of Ferrara, via Fossato di Mortara 17, I-44121 Ferrara, Italy. E-mail: [email protected]

b Department of Chemistry, University of Wroclaw, F. Joliot-Curie 14, 50-383 Wroclaw, Poland. E-mail: [email protected]

c Department of Chemistry and dIlse Katz Institute for Nanoscale Science and Technology, Ben Gurion University of the Negev, Beer-Sheva 84105, Israel. E-mail: [email protected]

Electronic Supplementary Material (ESI) for Dalton Transactions.This journal is © The Royal Society of Chemistry 2016

mailto:[email protected]

Table S1: Complex-formation constants for the ligand L1 with Cu(II), at T = 298 °K and I = 0.1 mol·dm−3 (KCl). Standard deviations on the last figure in parentheses.

Species Log pKa[CuLH3]2+ 25.68 (6) 4.3[CuLH2]+ 21.38 (3) 5.19[CuLH] 16.19 (4) 6.82[CuL]- 9.37 (6) 7.87

[CuLH-1]2- 1.50 (7) 8.25[CuLH-2]3- -6.75 (8) 10.05[CuLH-3]4- -16.8 (1) -

Table S2: Complex-formation constants for the ligand L2 with Cu(II), at T = 298 °K and I = 0.1 mol·dm−3 (KCl). Standard deviations on the last figure in parentheses.

Species Log pKa[CuLH4]3+ 32.26 (5) 4.44[CuLH3]2+ 27.82 (4) 4.58[CuLH2]+ 23.24 (3) 5.91[CuLH] 17.33 (4) 6.43[CuL]- 10.90 (5) -

[CuLH-2]3- -5.16 (6) 9.40[CuLH-3]4- -14.56 (8) -

Table S3: Spectroscopic parameters at different pH values for the system Cu(II)/L1; C°Cu(II) = 5·10−4 mol·dm−3; M/L ratio = 1.1.1.

UV-Vis CD EPR

pH Species /nm ε/M-1cm-1 λ/nm Δε/M-1 cm-1 AII (G) gII

4.5 [CuLH3]2+ [CuLH2]+ 663 41.1 254 0.15 164 2.32

5.0 [CuLH2]+ [CuLH] 635 67.2 580

250-0.131.36 166 2.28

5.6 [CuLH2]+ [CuLH] 622 80.8 586

252-0.212.17

6.2 [CuLH] [CuL]- 610 87.7 570245

-0.252.89 173 2.27

7.1 [CuL]- [CuLH] [CuLH-1]2- 608 90 570

250-0.253.11 186 2.25

7.5 [CuL]- [CuLH][CuLH-1]2- 603 92

677557457341250

0.10-0.300.06-0.133.47

8.0 [CuL]- [CuLH-1]2- [CuLH-2]3- 596 106

640548478336250

0.21-0.440.99-1.074.77

188 2.23

8.6 [CuLH-1]2- [CuLH-2]3- 577 123

633551474335252

0.44-0.460.16-1.725.41

9.1 [CuLH-1]2-

[CuLH-2]3- 562 140

638551474335254

0.51-0.460.16-1.725.59

188 2.23

9.6 [CuLH-2]3-

[CuLH-3]4- 558 130

635531402337254

0.80-0.260.16-1.245.96

10.3 [CuLH-2]3-

[CuLH-3]4- 530 128

631487402345260

0.96-0.700.06-0.536.12

186 2.22

11.2 [CuLH-3]4- 521 135

626486402318294258

1.37-1.180.020.76-0.196.78

193 2.19

Table S4: Spectroscopic parameters at different pH values for the system Cu(II)/L2; C°Cu(II) = 5·10−4 mol·dm−3; M/L ratio = 1.1.1.

UV-Vis CD EPR

pH Species /nm ε/M-1cm-1 λ/nm Δε/M-1 cm-1 AII (G) gII

4.7[CuLH4]3+ [CuLH3]2+ [CuLH2]+

659 36.1 252 0.63 164 2.31

5.2 [CuLH3]2+ [CuLH2]+ 639 52.0 249 1.30 169 2.28

6.2[CuLH2]+

[CuLH] [CuL]-

607 68.1 246 1.75 - -

6.7 [CuLH] [CuL]- 607 69.3 246 2.00 173 2.27

7.2 [CuL]- 602 73.4 247 2.44

7.7 [CuL]- [CuLH-2]3- 588 82.6

635557251

0.07-0.192.75

182 2.25

8.4 [CuL]- [CuLH-2]3- 579 89.8

636549476335247

0.20-0.390.09-1.214.41

186 2.21

8.8 [CuLH-2]3- [CuLH-3]4- 556 100.6

632548402336249

0.40-0.580.02-1.765.91

- -

10.0 [CuLH-2]3- [CuLH-3]4- 537 105.06

624517404339256

0.83-0.420.10-0.966.37

188 2.20

10.9 [CuLH-3]4- 522 108.5

624485353316294259

1.18-0.99-0.240.700.256.46

191 2.20

11.5 [CuLH-3]4- 518 109.3

626489358320294261

1.24-1.13-0.160.87-0.166.84

- -

Table S5: Metal-ligand interactions (L1a and L1b complexes).

X Cu-X [Å]L1a

His7 1.832His11 1.835

L1bHis5 2.440His7 1.849His11 1.841

Table S6: Metal-ligand interactions (L2 complex).

X Cu-X [Å]L2

His4 1.875His6 2.130His10 1.901

Table S7: Hydrogen bonds of L1a and L1b complexes. Fragments marked with star supply hydrogen bond with atoms form side chains.

Residue H..PA[Å]

PD-H..PA[deg]

Fragment HB type

L1aD2..H11 1.604 162.3 C=O*..H-N*D2..H11 1.637 170.6 C=O*..H-NA6..A8 2.195 157.8 N-H..O=C 3-10 helixA6..H9 2.105 151.3 C=O..H-N*Ac..D2 2.005 153.8 C=O(Ac)..H-NAc..D2 1.828 168.4 C=O(Ac)..H-N

L1bH5...A12 1.878 166.2 N-H*..O=CE1..H11 1.703 163.0 N-H..O=C*H7..H9 2.461 162.7 C=O..H-N 3-10 helixA6..A8 1.905 167.1 C=O..H-N 3-10 helixD2..D3 1.950 168.6 C=O*..H-NAc..A6 1.847 162.5 C=O(Ac)..H-N

Table S8: Hydrogen bonds of S2 complex. Fragments marked with star supply hydrogen bond with atoms form side chains.

Residue H..PA[Å]

PD-H..PA[deg]

Fragment HB type

S2E1..D3 1.987 150.7 N-H..O=C*D3..H4 1.180 153.6 C=O*..H-N*D3..A5 1.679 167.9 O=C..N-H 3-10 helixH4..H6 2.001 165.6 O=C..N-H 3-10 helixA5..A7 1.788 165.0 O=C..N-H 3-10 helixH6..H8 2.181 164.9 O=C..N-H 3-10 helixA7..A9 2.095 156.2 O=C..N-H 3-10 helix

A7..NH2 2.079 158.8 O=C..N-H2

H4..G13 1.704 162.3 N-H*..O=CH10..A11 1.670 154.2 N-H..O=C

Table S9: Conformational energies and populations obtained from the GBMV and Monte-Carlo calculations.

Peptide Averaged conformational energy

[kcal/mol]

Standard error of the averaged conformational energy values

[kcal/mol]

Populations (%)

L1a -819.9 0.7 27

L1b -861.0 0.6 73

2 3 4 5 6 7 8 90

10

20

30

40

50

60

70

80

90

100

% fo

rmat

ion

rela

tive

to [L

]

pH

L3-

LH3+6 LH2+

5

LH+4

LH3

LH-2

LH2-

LH4+7

Figure S1. Species distribution diagram for the protonation equilibria of the ligand L1; C°L = 6·10−4 mol·dm−3.

2 3 4 5 6 7 8 90

10

20

30

40

50

60

70

80

90

100%

form

atio

n re

lativ

e to

[L]

pH

LH5+8

LH4+7

LH3+6

LH2+5

LH+4

LH3

LH-2

LH2-

L3-

Figure S2. Species distribution diagram for the protonation equilibria of the ligand L2; C°L = 6·10−4 mol·dm−3.

460.5

562.3

604.8

648.2

690.3

721.2

879.2926.4

1087.3

200 400 600 800 1000 1200 1400 m/z0

2

4

6

9x10Intens.

Figure S3. ESI-MS spectrum of the system Cu(II)/L1; C°Cu(II) = 5·10−4 mol·dm−3; M/L ratio = 2.1; pH = 5; positive ion mode.

688.3

718.7

1377.5

1475.4

400 600 800 1000 1200 1400 1600 m/z0.0

0.5

1.0

1.5

2.0

2.5

8x10Intens.

Figure S4 ESI-MS spectrum of the system Cu(II)/L1; C°Cu(II) = 5·10−4 mol·dm−3; M/L ratio = 2.1; pH = 5; negative ion mode.

1440.5

1441.0

1441.5

1442.0

1442.5

1443.0

1443.5

1444.5

1445.51446.5

2014_04_04_Asia_Davide_CuL1_21_pos_do_kal_000001.d: +MS

1440.5

1441.5 1442.5

1443.5

1444.51445.5

2014_04_04_Asia_Davide_CuL1_21_pos_do_kal_000001.d: CuC56H77N22O20, M ,1440.500.00

0.25

0.50

0.75

1.00

1.25

1.508x10

Intens.

0

500

1000

1500

2000

2500

1440 1441 1442 1443 1444 1445 1446 1447 1448 m/z

Figure S5. Experimental (up) and simulated (down) ESI-MS spectrum for the complex [CuLH2]+ (CuC56H77N22O20, MW = 1440.6) in the system Cu(II)/L1; C°Cu(II) = 5·10−4 mol·dm−3; M/L ratio = 2.1; pH = 5; positive ion mode.

718.7 719.2

719.7

720.2

720.7721.2 721.7

2014_04_04_Asia_Davide_CuL1_21_low_neg_000001.d: -MS

718.7

719.2 719.7

720.2

720.7721.2

2014_04_04_Asia_Davide_CuL1_21_low_neg_000001.d: C56H74N22O20Cu, M ,1437.470.0

0.2

0.4

0.6

0.8

8x10Intens.

0

500

1000

1500

2000

2500

718.5 719.0 719.5 720.0 720.5 721.0 721.5 722.0 m/z

Figure S6. Experimental (up) and simulated (down) ESI-MS spectrum for the complex [CuLH-1]2- (CuC56H74N22O20, MW = 1437.6) in the system Cu(II)/L1; C°Cu(II) = 5·10−4 mol·dm−3; M/L ratio = 2.1; pH = 5; negative ion mode.

1500.41500.9

1501.4

1501.9

1502.4

1502.9

1503.4

1503.9

1504.4

1504.9

1505.4

1505.9

1506.41507.4

1508.4


1501.4

1502.4

1503.4

1504.4

1505.4

1506.4

1507.4

2014_04_04_Asia_Davide_CuL1_21_pos_do_kal_000001.d: C56H75N22O20Cu2, M ,1501.410

2

4

6

7x10Intens.

0

500

1000

1500

2000

2500

1500 1502 1504 1506 1508 1510 m/z

Figure S7. Experimental (up) and simulated (down) ESI-MS spectrum for the complex [Cu2L]+ (Cu2C56H75N22O20, MW = 1501.4) in the system Cu(II)/L1; C°Cu(II) = 5·10−4 mol·dm−3; M/L ratio = 2.1; pH = 5; positive ion mode.

524.5

590.2

637.8668.7

723.3

754.3

785.2

817.2

200 400 600 800 1000 1200 1400 m/z0

2

4

6

8

8x10Intens.

+MS

Figure S8. ESI-MS spectrum of the system Cu(II)/L2; C°Cu(II) = 5·10−4 mol·dm−3; M/L ratio = 2.1; pH = 5; positive ion mode.

401.7

721.3

1443.5

200 400 600 800 1000 1200 1400 1600 1800 2000 m/z0.0

0.5

1.0

1.5

8x10Intens.

Figure S9. ESI-MS spectrum of the system Cu(II)/L2; C°Cu(II) = 5·10−4 mol·dm−3; M/L ratio = 2.1; pH = 5; negative ion mode.

753.8754.3

754.8

755.3

755.8756.3

2014_03_25_Asia_Davide_Lig2Cu_pos_do_kal_000001.d: +MS

0.0

0.5

1.0

1.5

9x10Intens.

754.0 754.5 755.0 755.5 756.0 756.5 757.0 m/z

753.8

754.3 754.8

755.3

755.8756.3

2014_03_25_Asia_Davide_Lig2Cu_pos_medium_000001.d: C59H80N24O20Cu, M ,1507.53

0

500

1000

1500

2000

2500

754.0 754.5 755.0 755.5 756.0 756.5 757.0 m/z

Figure S10. Experimental (up) and simulated (down) ESI-MS spectrum for the complex [CuLH3]2+ (CuC59H80N24O20, MW = 1507.6) in the system Cu(II)/L2; C°Cu(II) = 5·10−4 mol·dm−3; M/L ratio = 2.1; pH = 5; positive ion mode.

803.2803.7

804.2804.7

805.2

805.4

805.7

806.2 806.7806.9

807.2

807.3

807.6


0.0

0.5

1.0

1.5

7x10Intens.

803.0 803.5 804.0 804.5 805.0 805.5 806.0 806.5 807.0 807.5 m/z

803.2

803.7

804.2

804.7

805.2

805.7

806.2

2014_03_25_Asia_Davide_Lig1Cu_pos_medium_000001.d: C59H77N24O20Cu2K, M ,1606.40

0

500

1000

1500

2000

2500

803.0 803.5 804.0 804.5 805.0 805.5 806.0 806.5 807.0 807.5 m/z

Figure S11. Experimental (up) and simulated (down) ESI-MS spectrum for the complex [Cu2L]K2+ (Cu2C59H77N24O20K, MW = 1606.4) in the system Cu(II)/L2; C°Cu(II) = 5·10−4 mol·dm−3; M/L ratio = 2.1; pH = 5; positive ion mode.

2500 2600 2700 2800 2900 3000 3100 3200 3300 3400 3500 3600 3700

Cu(II) - Ac-EDDAHAHAHAHAG-NH2

pH 8.18pH 6.14

pH 3.15

pH 5.15pH 4.16

pH 9.16

pH 11.11pH 10.10

Magnetic field [G]

pH 7.16

pH AII [G] gII

3.15 120.6 2.424.16 164.4 2.325.15 166.5 2.286.14 173.1 2.277.16 186.3 2.258.18 188.3 2.239.11 188.5 2.23

10.10 186.3 2.2211.11 192.8 2.19

Figure S12. EPR spectra at different pH for the system Cu(II)/L1; C°Cu(II) = 1·10−3 mol·dm−3; M/L ratio = 1.1.1.

2500 2600 2700 2800 2900 3000 3100 3200 3300 3400 3500 3600 3700

pH 10.50pH 9.50pH 8.67

pH 3.81pH4.77pH 5.50pH 6.67pH 7.40

Magnetic field [G]

pH 3.11

Cu(II) - Ac-EDDHAHAHAHAHG-NH2

pH A II[G] g II3.11 120.6 2.413.81 - 2.324.77 164.4 2.305.50 168.8 2.286.67 173.2 2.277.40 182.0 2.258.67 186.0 2.219.50 188.5 2.20

10.50 190.7 2.20

Figure S13. EPR spectra at different pH for the system Cu(II)/L2; C°Cu(II) = 1·10−3 mol·dm−3; M/L ratio = 1.1.1.

Figure S14 DSSP calculations for each residue along the peptides along the time of the simulations.

Figure S15 Populations of the simulated peptides L1a and L1b models. The populations had been estimated via Monte Carlo simulations.

Figure S16 The Cu(II)-Nε (His) atom and Cu(II)-O (carboxyl group) atoms distance distribution for each His residue obtained from MD simulations for the structures L1a, L1b and L2. The vertical lines within each box represent the median distance values.

L2

Figure S17. Synthetic network of hydrogen bonds in Complexes L1a, L1b and L2.

L1a

L1b

(a)

3 4 5 6 7 8 9 10 110

10

20

30

40

50

60

70

80

90

100

% for

matio

n rela

tive t

o [Cu

(II)]

pH

Cu(II) - Ac-THHHHAHGG-NH2


free Cu(II)

(b)

3 4 5 6 7 8 9 10 110

10

20

30

40

50

60

70

80

90

100

% fo

rmtio

n re

lative

to [C

u(II)

]

pH

free Cu(II)


Cu(II) - Ac-THHHHAHGG-NH2

Figure S18. Competition plot for a ternary solution containing equimolar concentrations (1 mM) of: (a) Cu(II), L1 and Ac- THHHHAHGG-NH2 (H. pylori); (b) Cu(II), L2 and Ac- THHHHAHGG-NH2 (H. pylori).

(a)

3 4 5 6 7 8 9 10 110

10

20

30

40

50

60

70

80

90

100

% fo

rmati

on re

ltive t

o [Cu

(II)]

pH

Cu(II) - Ac-PVHTGHMGHIGHTGHTGHTGSSGHG-NH2


free Cu(II)

(b)

3 4 5 6 7 8 9 10 110

10

20

30

40

50

60

70

80

90

100

Cu(II) - Ac-PVHTGHMGHIGHTGHTGHTGSSGHG-NH2

% fo

rmati

on re

lative

to [C

u(II)

]

pH


free Cu(II)

Figure S19. Competition plot for a ternary solution containing equimolar concentrations (1 mM) of: (a) Cu(II), L1 and Ac- PVHTGHMGHIGHTGHTGHTGHTGSSHG-NH2 (zp-PrP63-87); (b) Cu(II), L2 and Ac- PVHTGHMGHIGHTGHTGHTGHTGSSHG-NH2 (zp-PrP63-87).

(a)

3 4 5 6 7 8 9 10 110

10

20

30

40

50

60

70

80

90

100

% fo

rmat

ion re

lative

to [C

u(II)

]

pH

free Cu(II)

Cu(II) - Ac-HHHHHH-NH2


(b)

3 4 5 6 7 8 9 10 110

10

20

30

40

50

60

70

80

90

100


Cu(II) - Ac-HHHHHH-NH2

free Cu(II)

% fo

rmat

ion re

lative

to [C

u(II)

]

pH

Figure S20. Competition plot for a ternary solution containing equimolar concentrations (1 mM) of: (a) Cu(II), L1 and Ac-HHHHHH-NH2 (His6-tag) ; (b) Cu(II), L2 and Ac-HHHHHH-NH2 (His6-tag).

Documents

their mutated derivatives The unusual metal ion binding ability of … · 2016. 2. 17. · E-mail: [email protected] b Department of Chemistry, University of Wroclaw, F. Joliot-Curie 14,