TY - JOUR
T1 - Overcoming a hemihedral twinning problem in tetrahydrofolate-dependent O-demethylase crystals by the microseeding method
AU - Harada, Ayaka
AU - Sato, Yukari
AU - Kamimura, Naofumi
AU - Venugopalan, Nagarajan
AU - Masai, Eiji
AU - Senda, Toshiya
N1 - Funding Information:
This work was supported by Research Fellowships for Young Scientists from the Japan Society for the Promotion of Science (JSPS) and the 'Course-by-Course Education Program to Develop Student Research Capability and Aptitude of SOKENDAI to AH, and in part by JSPS KAKENHI 16K14908 to YS. GM/CA@APS is funded in whole or in part with federal funds from the National Cancer Institute (ACB- 12002) and the National Institute of General Medical Sciences (AGM-12006). This research used resources of the Advanced Photon Source, a US Department of Energy (DOE) Office of Science User Facility operated for the DOE Office of Science by Argonne National Laboratory under Contract No. DEAC02- 06CH11357.
Publisher Copyright:
© International Union of Crystallography, 2016.
PY - 2016/12/1
Y1 - 2016/12/1
N2 - A tetrahydrofolate-dependent O-demethylase, LigM, from Sphingobium sp. SYK-6 was crystallized by the hanging-drop vapour-diffusion method. However, the obtained P3121 or P3221 crystals, which diffracted to 2.5-3.3 Å resolution, were hemihedrally twinned. To overcome the twinning problem, microseeding using P3121/P3221 crystals as microseeds was performed with optimization of the reservoir conditions. As a result, another crystal form was obtained. The newly obtained crystal diffracted to 2.5-3.0 Å resolution and belonged to space group P21212, with unit-cell parameters a = 102.0, b = 117.3, c = 128.1 Å. The P21212 crystals diffracted to better than 2.0 Å resolution after optimizing the cryoconditions. Phasing using the single anomalous diffraction method was successful at 3.0 Å resolution with a Pt-derivative crystal. This experience suggested that microseeding is an effective method to overcome the twinning problem, even when twinned crystals are utilized as microseeds.A hemihedral twinning problem was found in crystals of a tetrahydrofolate-dependent O-demethylase, LigM, from Sphingobium sp. SYK-6. The problem was overcome using microseeding.
AB - A tetrahydrofolate-dependent O-demethylase, LigM, from Sphingobium sp. SYK-6 was crystallized by the hanging-drop vapour-diffusion method. However, the obtained P3121 or P3221 crystals, which diffracted to 2.5-3.3 Å resolution, were hemihedrally twinned. To overcome the twinning problem, microseeding using P3121/P3221 crystals as microseeds was performed with optimization of the reservoir conditions. As a result, another crystal form was obtained. The newly obtained crystal diffracted to 2.5-3.0 Å resolution and belonged to space group P21212, with unit-cell parameters a = 102.0, b = 117.3, c = 128.1 Å. The P21212 crystals diffracted to better than 2.0 Å resolution after optimizing the cryoconditions. Phasing using the single anomalous diffraction method was successful at 3.0 Å resolution with a Pt-derivative crystal. This experience suggested that microseeding is an effective method to overcome the twinning problem, even when twinned crystals are utilized as microseeds.A hemihedral twinning problem was found in crystals of a tetrahydrofolate-dependent O-demethylase, LigM, from Sphingobium sp. SYK-6. The problem was overcome using microseeding.
KW - O-demethylase
KW - Sphingobium sp. SYK-6
KW - cryoprotectant
KW - hemihedral twinning
KW - microseeding
KW - tetrahydrofolate
UR - http://www.scopus.com/inward/record.url?scp=85002496981&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=85002496981&partnerID=8YFLogxK
U2 - 10.1107/S2053230X16018665
DO - 10.1107/S2053230X16018665
M3 - Article
C2 - 27917838
AN - SCOPUS:85002496981
SN - 1744-3091
VL - 72
SP - 897
EP - 902
JO - Acta Crystallographica Section F:Structural Biology Communications
JF - Acta Crystallographica Section F:Structural Biology Communications
IS - 12
ER -