Structural basis for the specific inhibition of protein kinase G, a virulence factor of Mycobacterium tuberculosis

Area of Science:

• Microbiology
• Structural Biology
• Biochemistry

Background:

• Pathogenic mycobacteria, including Mycobacterium tuberculosis, survive within host macrophages by utilizing virulence factors.
• Protein kinase G (PknG) is a key virulence factor that prevents the degradation of mycobacteria in lysosomes.
• Inhibiting PknG with AX20017 leads to lysosomal targeting and killing of mycobacteria.

Purpose of the Study:

• To elucidate the structural basis of PknG's function and its inhibition by AX20017.
• To understand the unique structural features of PknG that enable selective targeting.
• To provide insights into developing novel anti-tubercular therapies.

Main Methods:

• X-ray crystallography was used to determine the structure of PknG in complex with AX20017 at 2.4 Å resolution.
• Directed mutagenesis was employed to investigate the roles of different PknG domains and specific residues.
• Biochemical assays were used to assess the inhibitory potency of AX20017 and the impact of mutations.

Main Results:

• The crystal structure revealed a unique multidomain topology of PknG, comprising N-terminal rubredoxin, central kinase, and C-terminal tetratrico-peptide repeat domains.
• The rubredoxin domain was identified as a regulator of PknG kinase activity.
• AX20017 binds to a unique adenosine-binding pocket in an active conformation of the kinase domain, distinct from human kinases.
• Mutagenesis of unique residues in the binding pocket abolished AX20017's inhibitory potency.

Conclusions:

• The study explains the specific mechanism of action of AX20017 against PknG.
• The findings demonstrate that targeting mycobacterial virulence factors, even those homologous to host molecules, is a viable strategy to combat Mycobacterium tuberculosis proliferation.
• The unique structural features of PknG offer opportunities for developing highly selective inhibitors.