Sulphate removal induces a major conformational change in Leishmania mexicana pyruvate kinase in the crystalline state

Lindsay B. Tulloch, Hugh P. Morgan, Véronique Hannaert, Paul A. M. Michels, Linda A. Fothergill-Gilmore, Malcolm D. Walkinshaw (Lead / Corresponding author)

Research output: Contribution to journalArticlepeer-review

22 Citations (Scopus)


We report X-ray structures of pyruvate kinase from Leishmania mexicana (LmPYK) that are trapped in different conformations. These, together with the previously reported structure of LmPYK in its inactive (T-state) conformation, allow comparisons of three different conformers of the same species of pyruvate kinase (PYK). Four new site point mutants showing the effects of side-chain alteration at subunit interfaces are also enzymatically characterised. The LmPYK tetramer crystals grown with ammonium sulphate as precipitant adopt an active-like conformation, with sulphate ions at the active and effector sites. The sulphates occupy positions similar to those of the phosphates of ligands bound to active (R-state) and constitutively active (nonallosteric) PYKs from several species, and provide insight into the structural roles of the phosphates of the substrates and effectors. Crystal soaking in sulphate-free buffers was found to induce major conformational changes in the tetramer. In particular, the unwinding of the Aalpha6' helix and the inward hinge movement of the B domain are coupled with a significant widening (4 A) of the tetramer caused by lateral movement of the C domains. The two new LmPYK structures and the activity studies of site point mutations described in this article are consistent with a developing picture of allosteric activity in which localised changes in protein flexibility govern the distribution of conformer families adopted by the tetramer in its active and inactive states.

Original languageEnglish
Pages (from-to)615-626
Number of pages12
JournalJournal of Molecular Biology
Issue number3
Early online date23 Aug 2008
Publication statusPublished - 14 Nov 2008


  • Adenosine Triphosphate/chemistry
  • Allosteric Regulation
  • Amino Acid Sequence
  • Animals
  • Binding Sites
  • Crystallography, X-Ray
  • Enzyme Activation
  • Humans
  • Leishmania mexicana/enzymology
  • Models, Molecular
  • Molecular Sequence Data
  • Mutagenesis, Site-Directed
  • Point Mutation
  • Protein Structure, Quaternary
  • Pyruvate Kinase/chemistry
  • Sulfates/chemistry
  • conformational transitions
  • Leishmania mexicana
  • pyruvate kinase
  • X-ray crystallography


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