AMPK beta subunits display isoform specific affinities for carbohydrates

Area of Science:

• Biochemistry
• Molecular Biology
• Structural Biology

Background:

• AMP-activated protein kinase (AMPK) is a crucial cellular energy sensor, composed of alpha, beta, and gamma subunits.
• Both beta isoforms (beta1 and beta2) possess a mid-molecule carbohydrate-binding module (beta-CBM).
• AMPK beta2 is predominantly expressed in muscle tissue, suggesting specialized roles.

Purpose of the Study:

• To investigate the structural and functional evolution of the AMPK beta-CBM, particularly the beta2 isoform.
• To determine the role of a specific threonine (Thr) insertion in the beta2-CBM's function.
• To elucidate the binding preferences of beta-CBM for specific carbohydrate structures.

Main Methods:

• Comparative sequence analysis of beta-CBM across species.
• Biochemical assays to measure binding affinity to various oligosaccharides and glycogen mimetics.
• Site-directed mutagenesis to introduce or delete the Thr residue in beta-CBM.

Main Results:

• The beta2-CBM has evolved a unique Thr insertion compared to beta1-CBM.
• This Thr insertion significantly increases the beta2-CBM's affinity for oligosaccharides with a single alpha-1,6 branched residue.
• Deletion of Thr-101 in beta2-CBM reduced affinity by 3-fold; its insertion into beta1-CBM increased affinity by 3-fold.

Conclusions:

• The Thr insertion in AMPK beta2-CBM is a key evolutionary adaptation enhancing its interaction with specific glycogen-like structures.
• This finding highlights the functional importance of specific residues in regulating substrate binding and potentially AMPK activity in muscle.
• The study provides insights into the molecular mechanisms underlying AMPK regulation and its role in energy metabolism.