Allostery and conformational dynamics in cAMP-binding acyltransferases.

Area of Science:

• Biochemistry and Molecular Biology
• Structural Biology
• Microbial Pathogenesis

Background:

• Mycobacteria possess unique proteins, lysine acyltransferases (KATs), that control protein lysine acylation.
• KATs from Mycobacterium smegmatis (KATms) and Mycobacterium tuberculosis (KATmt) exhibit distinct biochemical characteristics.
• Differences lie in cyclic adenosine monophosphate (cAMP) binding affinity and allosteric activation of the acyltransferase domain.

Purpose of the Study:

• To elucidate the structural basis for KATms' high cAMP binding affinity and activity.
• To understand the divergence in biochemical properties between KATms and KATmt.
• To identify the mechanisms responsible for altered protein conformational states and enzymatic activity.

Main Methods:

• Structure-guided mutational analysis of KATms.
• Biochemical assays to assess cAMP binding and acyltransferase activity.
• Comparative analysis of amino acid sequences and structural features.

Main Results:

• Identified specific structural features in KATms responsible for high affinity cAMP binding.
• Demonstrated that KATms exhibits elevated acyltransferase activity even without cAMP.
• Mutational analysis converted KATms into a cAMP-dependent enzyme and revealed a unique asparagine residue in KATms aiding catalysis.

Conclusions:

• Divergence in KATms and KATmt properties is driven by distinct amino acid sequences.
• A unique asparagine residue in KATms facilitates enzymatic activity independently of a conserved glutamate found in related enzymes.
• Amino acid modifications significantly alter protein conformational states, impacting enzymatic regulation in mycobacteria.