An electron microscopic study of pyruvate carboxylase

Area of Science:

• Biochemistry
• Enzymology
• Structural Biology

Background:

• Pyruvate carboxylase is a crucial enzyme in gluconeogenesis and anaplerosis.
• Previous structural studies on vertebrate pyruvate carboxylases yielded conflicting results, with some reporting a square-planar tetrameric structure.

Purpose of the Study:

• To elucidate the native structure of pyruvate carboxylase from vertebrate sources (sheep and chicken liver).
• To compare the structure of vertebrate pyruvate carboxylase with that of yeast pyruvate carboxylase.
• To resolve discrepancies in previous structural characterizations of vertebrate pyruvate carboxylase.

Main Methods:

• Purification of pyruvate carboxylase to homogeneity from sheep and chicken liver.
• Electron microscopy for high-resolution structural analysis of the purified enzymes.

Main Results:

• Pyruvate carboxylases from sheep and chicken liver exhibit a splayed tetrahedral structure.
• This structure, in certain electron microscopic views, resembles the reported structure of yeast pyruvate carboxylase.
• The previously described square-planar tetrameric structure was attributed to a larger, biotin-devoid molecule, not pyruvate carboxylase.

Conclusions:

• The native structure of vertebrate pyruvate carboxylase is splayed tetrahedral, not square-planar.
• The presence of biotin is essential for the correct structure and function of pyruvate carboxylase.
• Clarification of pyruvate carboxylase structure aids understanding of its role in metabolic pathways.