Han GW et al. “Crystal structure of a metal-dependent phosphoesterase (YP_910028.1) from Bifidobacterium
adolescentis: Computational prediction and experimental validation of phosphoesterase activity”
Supplementary Materials
Identification of metal ions
X-ray fluorescence emission peaks for selenium, zinc and iron were observed when the
crystal was excited with X-rays 500 eV above the Se-edge at SSRL beamline 1-5. To
determine the identity of the metals at the individual sites in the structure, four additional
data sets were collected on SSRL beamline 9-2. These datasets were collected to 3-3.5Å
Å resolution at wavelengths 1.276 Å, 1.293 Å, 1.729 Å and 1.756 Å, which are above
and below the zinc and iron absorption edges. (The K absorption edges of Zn and Fe
atoms are 9659 eV and 7112 eV, respectively.) The data were integrated with MOSFLM
and scaled with the program SCALA. Data statistics are described in Table S1.
Anomalous difference Fourier maps were calculated for each wavelength using the final
model phase (retarded by 90°) and anomalous difference amplitudes. The large changes
in peak heights across the zinc or iron absorption edge clearly showed that one site was
zinc and the other two were iron. The integrated peak heights at the metal sites are
shown with the peak heights for selenium and sulfur sites as a reference in Table S2. The
theoretical f” values at each wavelength are listed for comparison in Table S3.
Table S1 – Data collection statistics for metal site identification from the ApoYP_910028.1 crystal.
Data collection
Wavelength (Å)
Resolution range (Å)
Number of observations
Number of unique
reflections
Completeness(%)
Mean I/σ(I)
Rmerge on I (%)
Highest resolution shell (Å)
λ2 above Zn
1.2755
28.7-3.00
24,541
6,755
λ3 below Zn
1.2928
28.7-3.08
22,652
6,267
λ4 above Fe
1.7291
28.7-3.46
15,460
3,460
λ5 below Fe
1.7561
28.7-3.46
15,457
4,432
99.9 (100.0)
8.3 (2.0)
15.6 (56.6)
3.08-3.00
99.9 (100.0)
8.1 (1.7)
16.5 (61.8)
3.16-3.08
99.8 (100.0)
6.3 (1.4)
23.3 (75.0)
3.55-3.46
99.8 (100.0)
6.2 (1.5)
21.8 (69.7)
3.55-3.46
Highest resolution shell is shown in parentheses.
Rmerge= ΣhklΣi | Ii(hkl) - <I(hkl)> | / ΣhklΣi Ii(hkl)
Table S2 - Anomalous difference Fourier peaks from the Zn and Fe atoms.
Zn-301
Fe-302
Fe-303
Selenium (Se) Sulfur (Sγ) of
of Mse 116
Cys 18
λ2 above Zn (9720 eV)
3.57
3.22
3.01
1.30
0.93
λ3 below Zn (9590 eV)
0.68
3.02
3.68
1.38
0.61
λ4 above Fe (7170 eV)
0.23
4.64
7.48
2.33
1.17
λ5 below Fe (7060 eV)
0.79
1.85
1.55
0.39
1.27
The signal listed is the value reported by MAPMAN after integration of a sphere of
radius 2.5 Å around the atom center from the final refined model. The selenium (Mse
116) and sulfur (Cys 18) values are included to provide a reference for differences in
scale between the four maps.
Table S3 – Theoretical f” values at each wavelength╫.
Data Set
Wavelength (Å)
Energy (eV)
Theoretical f” Se
Theoretical f” Zn
Theoretical f” Fe
Theoretical f” S
λ1 SADSe
0.9785
12670
3.84
2.47
1.50
0.23
λ2 above Zn
1.2755
9720
0.81
3.86
2.36
0.39
λ3 below Zn
1.2928
9590
0.83
0.49
2.39
0.40
λ4 above Fe
1.7291
7170
1.39
0.83
3.89
0.69
Theoretical f” data were extracted from the tables compiled by Ethan Merritt
(http://skuld.bmsc.washington.edu/scatter/AS_periodic.html)
╫
λ5 below Fe
1.7561
7060
1.43
0.86
0.48
0.71