Perdeuteration, large crystal growth and neutron data collection of Leishmania mexicana triose-phosphate isomerase E65Q variant

Vinardas Kelpšas, Bénédicte Lafumat, Matthew P. Blakeley, Nicolas Coquelle, Esko Oksanen, Claes Von Wachenfeldt

Research output: Contribution to journalArticlepeer-review

5 Citations (SciVal)

Abstract

Triose-phosphate isomerase (TIM) catalyses the interconversion of dihydroxyacetone phosphate and glyceraldehyde 3-phosphate. Two catalytic mechanisms have been proposed based on two reaction-intermediate analogues, 2-phosphoglycolate (2PG) and phosphoglycolohydroxamate (PGH), that have been used as mimics of the cis-enediol(ate) intermediate in several studies of TIM. The protonation states that are critical for the mechanistic interpretation of these structures are generally not visible in the X-ray structures. To resolve these questions, it is necessary to determine the hydrogen positions using neutron crystallography. Neutron crystallography requires large crystals and benefits from replacing all hydrogens with deuterium. Leishmania mexicana triose-phosphate isomerase was therefore perdeuterated and large crystals with 2PG and PGH were produced. Neutron diffraction data collected from two crystals with different volumes highlighted the importance of crystal volume, as smaller crystals required longer exposures and resulted in overall worse statistics.

Original languageEnglish
Pages (from-to)260-269
Number of pages10
JournalActa crystallographica. Section F, Structural biology communications
Volume75
Issue number4
DOIs
Publication statusPublished - 2019

Subject classification (UKÄ)

  • Structural Biology

Keywords

  • large crystals
  • Laue method
  • neutron diffraction
  • perdeuteration
  • triose-phosphate isomerase

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