Molecular and evolutionary determinants for protein interaction within a class II aldolase/Adducin domain

Area of Science:

• Evolutionary biology
• Molecular biology
• Structural biology

Background:

• Modular protein interaction domains are key to multicellularity.
• Class II aldolase domain (ALDODOM) in Adducin family resembles ancient glycolytic enzymes.
• ALDODOM typically binds actin filaments but its functional switch is unclear.

Purpose of the Study:

• Investigate the molecular basis of ALDODOM interaction with the mitotic spindle regulator, Mud.
• Elucidate the sequence changes underlying the functional switch of the aldolase protein fold.

Main Methods:

• Explored molecular interactions between Hts ALDODOM (Drosophila Adducin) and Mud.
• Utilized sequence database analyses to trace ALDODOM evolutionary origins.
• Performed mutational analysis, including C-terminal helix truncation and chimeric fusions.

Main Results:

• Conserved actin-binding residues are not essential for Mud binding.
• The ALDODOM C-terminal helix (CThelix) and adjacent protomer residues are critical for Mud interaction.
• Truncating CThelix or fusing it to bacterial aldolase conferred Mud binding.
• A single arginine-to-glycine change in bacterial aldolase also enabled Mud binding.
• ALDODOM function likely originated in early metazoans like Placozoa.

Conclusions:

• ALDODOM interaction with Mud involves a distinct mechanism from actin binding.
• The C-terminal helix is a key determinant of ALDODOM's interaction with Mud.
• Evolutionary analysis suggests ALDODOM's functional diversification occurred early in metazoan evolution.