TY - JOUR
T1 - Functional conformations for pyruvate carboxylase during catalysis explored by cryoelectron microscopy
AU - Lasso, Gorka
AU - Yu, Linda P.C.
AU - Gil, David
AU - Lázaro, Melisa
AU - Tong, Liang
AU - Valle, Mikel
N1 - Funding Information:
This work was supported by grant BFU2012-34873 from the Spanish Ministry of Economy and Competitiveness (to M.V.) and a grant from the NIH (DK067238 to L.T.). L.P.C.Y. was also supported by an NIH training program in Cellular and Molecular Foundations of Biomedical Science (GM008798).
PY - 2014/6/10
Y1 - 2014/6/10
N2 - Summary The tetrameric enzyme pyruvate carboxylase (PC), a biotin-dependent carboxylase, produces oxaloacetate by two consecutive reactions that take place in distant active sites. Previous crystal structures revealed two different configurations for PC tetramers, the so-called symmetric and asymmetric, which were understood as characteristic molecular architectures for PC from different organisms. We have analyzed PC samples from Staphylococcus aureus while the enzyme generates oxaloacetate, expecting PC tetramers to display the conformational landscape relevant for its functioning. Using cryoelectron microscopy (cryo-EM) and sorting techniques, we detect previously defined symmetric and asymmetric architectures, demonstrating that PC maps both arrangements by large conformational changes. Furthermore, we observe that each configuration is coupled to one of the two consecutive enzymatic reactions. The findings describe the structural transitions relevant for the allosteric control of the multifunctional PC and demonstrate that by cryo-EM and classification, we can characterize freely working macromolecules.
AB - Summary The tetrameric enzyme pyruvate carboxylase (PC), a biotin-dependent carboxylase, produces oxaloacetate by two consecutive reactions that take place in distant active sites. Previous crystal structures revealed two different configurations for PC tetramers, the so-called symmetric and asymmetric, which were understood as characteristic molecular architectures for PC from different organisms. We have analyzed PC samples from Staphylococcus aureus while the enzyme generates oxaloacetate, expecting PC tetramers to display the conformational landscape relevant for its functioning. Using cryoelectron microscopy (cryo-EM) and sorting techniques, we detect previously defined symmetric and asymmetric architectures, demonstrating that PC maps both arrangements by large conformational changes. Furthermore, we observe that each configuration is coupled to one of the two consecutive enzymatic reactions. The findings describe the structural transitions relevant for the allosteric control of the multifunctional PC and demonstrate that by cryo-EM and classification, we can characterize freely working macromolecules.
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U2 - 10.1016/j.str.2014.04.011
DO - 10.1016/j.str.2014.04.011
M3 - Article
C2 - 24882745
AN - SCOPUS:84902343204
SN - 0969-2126
VL - 22
SP - 911
EP - 922
JO - Structure
JF - Structure
IS - 6
ER -