Interdomain Conformational Changes Provide Allosteric Regulation en Route to Chorismate

Area of Science:

• Biochemistry
• Enzymology
• Structural Biology

Background:

• Multifunctional proteins are crucial for metabolic regulation.
• 3-deoxy-d-arabino heptulosonate-7-phosphate synthase (DAH7PS) initiates aromatic metabolite biosynthesis.
• Chorismate mutase (CM) directs the pathway towards tyrosine and phenylalanine synthesis.

Purpose of the Study:

• To investigate the catalytic functions and allosteric regulation of a combined DAH7PS-CM enzyme from Geobacillus sp.
• To elucidate the molecular mechanism underlying prephenate-mediated inhibition of DAH7PS activity.

Main Methods:

• Protein expression and purification of the full-length DAH7PS-CM fusion protein.
• Enzyme activity assays to assess catalytic function and inhibition.
• X-ray crystallography and small-angle X-ray scattering (SAXS) to determine protein structure and conformational changes.

Main Results:

• DAH7PS and CM activities reside in distinct, separable domains.
• Prephenate, the CM product, allosterically inhibits DAH7PS activity in the full-length fusion protein.
• Structural data reveals that prephenate binding induces a conformational change, increasing inter-domain association and occluding the DAH7PS active site.

Conclusions:

• Gene fusion creates a physical gating mechanism for allosteric control of DAH7PS activity by CM.
• This mechanism represents a general strategy for allosteric regulation in multifunctional enzymes.
• Understanding this allosteric regulation provides insights into metabolic pathway control.