Table 1

Summary of data collection and MAD structure determination. Two seleno-methionated FimC-FimH crystals (space groupC2, a = 139.1 Å, b = 139.1 Å, c = 214.5 Å, β = 90.0 Å) exhibiting strong pseudo P41212 symmetry were used to collect MAD (22) data on BM14 of the European Synchrotron Radiation Facility. Data were recorded at each of three wavelengths corresponding to the peak of the Se white line, the point of inflexion of the K absorption edge, and a remote wavelength by using a MAR charge-coupled device detector. Data were reduced with the program HKL2000 (23), with further processing and scaling using the CCP4 processing package (23). An initial solution to the Patterson function was produced in the tetragonal pseudo space group both automatically with the program SOLVE (23) and manually with the program RSPS (23), and initial phases were calculated with SHARP (23). Density modification including fourfold noncrystallographic (NCS) averaging was done with the program DM (23). A model corresponding to the two copies of the complex in the pseudo asymmetric unit was built with the program O (23). Bulk solvent correction, positional, simulated annealing, and isotropic temperature factor refinement was carried out with X-PLOR (23) and REFMAC (23) with tight NCS restraints against a 2.5 Å native data set collected at Max II/BL711 in Lund. The current R factor andR free (on 5% of the data) are 24.0 and 26.8%, respectively. The root mean square deviations from ideal bond lengths and angles are 0.016 and 3.3 Å, respectively. No residues are in disallowed regions of the Ramachandran plot.

Data collection statistics
Crystaldmin(Å)NuniqueCmplt*(%)MultI/σ(I)Rsym§(%)Ranom| (%)
SeMet Crystal 12.882.8
Remote93,0192.513.1 (3.7)4.0 (17.3)3.5 (16.8)
Point of inflection75,4672.111.6 (6.9)3.5 (24.4)4.3 (21.4)
Peak82,7542.711.3 (1.9)4.1 (24.7)4.2 (18.8)
SeMet Crystal 22.798.7
Remote110,9283.88.9 (2.0)5.1 (28.3)4.2 (20.9)
Point of inflection110,4154.010.6 (2.7)4.2 (21.8)3.8 (17.4)
Peak110,4183.914.4 (2.8)4.2 (20.8)4.2 (17.5)
Native2.5139,64598.04.15.3 (1.6)7.6 (25.3)NA
Phasing statistics from SHARP
Point of inflexion λ = 0.9793 ÅPeak λ = 0.9792 ÅRemote λ = 0.885 Å
CentricAcentricCentricAcentricCentricAcentric
Phasing power 2.0/−2.1/1.22.0/−2.0/1.6−/−−/0.81
R cullis # 0.49/−0.56/0.520.53/−0.54/0.57−/−−/0.69
Resolution (Å)7.595.504.523.933.533.232.992.80
FOM** 0.6230.5080.3790.2270.1720.1400.1050.125
  • * Completeness.

  • Multiplicity.

  • Overall value and values in parentheses are for the highest resolution shell.

  • § Rsym = ΣhΣi|Ii(h) − 〈I(h)〉|/ΣhΣiIi(h), where Ii(h) and 〈I(h)〉 are the intensities of the individual and mean structure factors, respectively. The high-resolution shell is in parentheses.

  • | Ranom = ΣhΣi|Ii(h) − 〈I(h)〉|/ΣhΣiIi(h);Ii(h) and 〈I(h)〉 are as defined above, and the summation is over anomalous pairs. The high-resolution shell is in parentheses.

  • FH(calc)/E, where E is the estimated lack-of-closure error (isomorphous/anomalous).

  • # Rcullis = Σ||FPH − FP| − FH(calc)|/Σ|FPH − FP|, ]where FP and FPH are protein and heavy-atom structure factors, respectively, andFH(calc) is the calculated heavy-atom structure factor (isomorphous/anomalous).

  • ** Figure of merit for SHARP phases.