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Figure 1
Characterization of phase errors for projection data. Phase errors are the best tool for the evaluation of data quality in electron crystallographic work. Shown in this figure is how different types of overall mean phase errors of projection data behave as a function of the number of images used. The errors were obtained by sequential averaging of data from 1, 2, 3, 4 and 20 images of a gap-junction channel (black columns; Unger, Kumar et al., 1997BB27) and the bacterial Na+/H+-antiporter NhaA (grey columns; Williams et al., 1999BB35). Data from the first three images were of truly untilted crystals. In all other cases, only those measurements within 0.0016 Å−1 (gap junction) and 0.005 Å−1 (NhaA) along the direction of z* were included in the averaging. Errors shown in (a) correspond to the overall figure of merit (FOM) calculated by AVRGAMPHS. An error of 90° would be obtained for random data. The errors shown in (b) represent the mean centric phase error. This error reflects the average deviation of the average phases from their theoretical target values (0/180°) imposed by the p6 (gap junctions) and p22121 (NhaA) symmetries of the two specimens. In this case, random data would result in a 45° deviation overall. The total number of independent measurements to 6 Å resolution was 179, 353, 516, 596 and 1266 in case of the gap junction, describing 76 unique reflections. In the case of NhaA, 368, 713, 936, 1152 and 1704 individual measurements were averaged to yield phases for 204 unique reflections. The smaller redundancy in the data and the lower symmetry in the case of NhaA explain the slightly higher phase errors shown in (a) compared with those obtained for the gap-junction specimen. However, the overall centric phase errors are very similar in both cases and do not improve much as data are added. This unexpected behavior is related to the 3D molecular transforms, which in both cases cause systematic errors in the determination of the projection data. The conclusion to be drawn here is that the use of a large and highly redundant data set will provide highly significant averages for the phase values. However, significance of the averages and their conformity with applicable phase constraints are not necessarily related to each other.

Journal logoBIOLOGICAL
ISSN: 1399-0047
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