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2016 ; 72
(Pt 3
): 421-9
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Data-collection strategy for challenging native SAD phasing
#MMPMID26960129
Olieric V
; Weinert T
; Finke AD
; Anders C
; Li D
; Olieric N
; Borca CN
; Steinmetz MO
; Caffrey M
; Jinek M
; Wang M
Acta Crystallogr D Struct Biol
2016[Mar]; 72
(Pt 3
): 421-9
PMID26960129
show ga
Recent improvements in data-collection strategies have pushed the limits of
native SAD (single-wavelength anomalous diffraction) phasing, a method that uses
the weak anomalous signal of light elements naturally present in macromolecules.
These involve the merging of multiple data sets from either multiple crystals or
from a single crystal collected in multiple orientations at a low X-ray dose.
Both approaches yield data of high multiplicity while minimizing radiation damage
and systematic error, thus ensuring accurate measurements of the anomalous
differences. Here, the combined use of these two strategies is described to solve
cases of native SAD phasing that were particular challenges: the integral
membrane diacylglycerol kinase (DgkA) with a low Bijvoet ratio of 1% and the
large 200 kDa complex of the CRISPR-associated endonuclease (Cas9) bound to guide
RNA and target DNA crystallized in the low-symmetry space group C2. The optimal
native SAD data-collection strategy based on systematic measurements performed on
the 266 kDa multiprotein/multiligand tubulin complex is discussed.
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