Sulphate removal induces a major conformational change in Leishmania mexicana pyruvate kinase in the crystalline state.
J Mol Biol
; 383(3): 615-26, 2008 Nov 14.
Article
in En
| MEDLINE
| ID: mdl-18775437
ABSTRACT
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.
Full text:
1
Collection:
01-internacional
Database:
MEDLINE
Main subject:
Pyruvate Kinase
/
Sulfates
/
Leishmania mexicana
/
Protein Structure, Quaternary
Limits:
Animals
/
Humans
Country/Region as subject:
Mexico
Language:
En
Journal:
J Mol Biol
Year:
2008
Document type:
Article
Affiliation country: