Mammalian fatty acid synthetase is a structurally and functionally symmetrical dimer

Area of Science:

• Biochemistry
• Enzymology
• Molecular Biology

Background:

• Mammalian fatty acid synthetase (FAS) is a multi-domain enzyme crucial for lipogenesis.
• Understanding the functional symmetry of its active sites is key to comprehending its catalytic mechanism.

Purpose of the Study:

• To experimentally determine if the two condensing-enzyme active centers of mammalian fatty acid synthetase are simultaneously functional.
• To investigate the structural symmetry and catalytic activity of the enzyme's core polypeptides.

Main Methods:

• Utilized trypsinized fatty acid synthetase (nicked homodimer) lacking thioesterase domains.
• Quantified 4'-phosphopantetheine content and long-chain acyl-enzyme synthesis using radiolabeled precursors.
• Employed cross-linking with 1,3-dibromo-2-propanone to probe active site accessibility and function.

Main Results:

• Stoichiometry indicated nearly two acyl chains assembled per holoenzyme dimer, suggesting full active site occupancy.
• Cross-linking experiments demonstrated that both condensing sites are accessible and functional.
• Inhibition of activity correlated with cross-linking, supporting a symmetrical model where active sites are juxtaposed.

Conclusions:

• The mammalian fatty acid synthetase exhibits a structurally and functionally symmetrical model.
• Both condensing-enzyme active centers are simultaneously active, contributing to efficient long-chain fatty acid synthesis.