Macromolecular impurities and disorder in protein crystals.
The mechanisms by which macromolecular impurities degrade the diffraction properties of protein crystals have been investigated using X-ray topography, high-resolution diffraction line shape measurements, crystallographic data collection, chemical analysis, and two-photon excitation fluorescence microscopy. Hen egg-white lysozyme crystals grown from solutions containing a structurally unrelated protein (ovotransferrin) and a related protein (turkey egg-white lysozyme) can exhibit significantly broadened mosaicity due to formation of cracks and dislocations but have overall B factors and diffraction resolutions comparable to those of crystals grown from uncontaminated lysozyme. Direct fluorescence imaging of the three-dimensional impurity distribution shows that impurities incorporate with different densities in sectors formed by growth on different crystal faces, and that impurity densities in the crystal core and along boundaries between growth sectors can be much larger than in other parts of the crystal. These nonuniformities create stresses that drive formation of the defects responsible for the mosaic broadening. Our results provide a rationale for the use of seeding to obtain high-quality crystals from heavily contaminated solutions and have implications for the use of crystallization for protein purification. Proteins 1999;36:270-281.
Caylor
C L
CL
Laboratory of Atomic and Solid State Physics, Cornell University, Ithaca, New York 14853, USA.
Dobrianov
I
I
Lemay
S G
SG
Kimmer
C
C
Kriminski
S
S
Finkelstein
K D
KD
Zipfel
W
W
Webb
W W
WW
Thomas
B R
BR
Chernov
A A
AA
Thorne
R E
RE
D000818:Animals; D002645:Chickens; D003207:Conalbumin / Q000302:isolation & purification; D003460:Crystallization; D018360:Crystallography, X-Ray; D004340:Drug Contamination; D046911:Macromolecular Substances; D008856:Microscopy, Fluorescence; D009113:Muramidase / Q000302:isolation & purification / Q000592:standards; D011506:Proteins / Q000302:isolation & purification* / Q000592:standards; D011786:Quality Control; D014422:Turkeys
D046911:Macromolecular Substances; D011506:Proteins; D003207:Conalbumin; D009113:Muramidase