Overcoming a hemihedral twinning problem in tetrahydrofolate-dependent O-demethylase crystals by the microseeding method

Ayaka Harada, Yukari Sato, Naofumi Kamimura, Nagarajan Venugopalan, Eiji Masai, Toshiya Senda

Research output: Contribution to journalArticle

1 Citation (Scopus)

Abstract

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.

Original languageEnglish
Pages (from-to)897-902
Number of pages6
JournalActa Crystallographica Section:F Structural Biology Communications
Volume72
Issue number12
DOIs
Publication statusPublished - 2016 Dec 1
Externally publishedYes

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Keywords

  • cryoprotectant
  • hemihedral twinning
  • microseeding
  • O-demethylase
  • Sphingobium sp. SYK-6
  • tetrahydrofolate

ASJC Scopus subject areas

  • Biophysics
  • Structural Biology
  • Biochemistry
  • Genetics
  • Condensed Matter Physics

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