Homodimerization through coiled-coil regions enhances activity of the myotonic dystrophy protein kinase

Area of Science:

• Biochemistry
• Molecular Biology
• Genetics

Background:

• Myotonic dystrophy protein kinase (DMPK) is linked to cardiac and behavioral issues in myotonic dystrophy.
• DMPK has distinct regions: leucine-rich repeat (L), kinase (PK), coiled-coil (H), and transmembrane tail (T).
• Homology exists between DMPK's PK and H regions and other protein kinases involved in cytoskeletal and cell cycle functions.

Purpose of the Study:

• To investigate the role of DMPK subfragments in dimerization.
• To determine the structural requirements for DMPK dimerization.
• To assess the impact of dimerization on DMPK's kinase activity.

Main Methods:

• Analysis of DMPK subfragments (LPKH, PKH, LPK) for dimerization.
• Investigating the effect of caspase-1 cleavage on LPKH dimers.
• Enzyme kinetics assays (Vmax, Km, kcat/Km) using a synthetic peptide substrate.

Main Results:

• LPKH and PKH subfragments formed dimers, while LPK remained monomeric, indicating the H domain is essential for dimerization.
• Caspase-1 cleavage of LPKH between PK and H domains resulted in monomeric forms, confirming the H domain's role in dimerization.
• Dimeric LPKH exhibited over 10-fold greater catalytic efficiency (Vmax and kcat/Km) compared to monomeric forms.
• Dimeric LPKH had a higher Km (over 3-fold) for the substrate, suggesting altered substrate binding.

Conclusions:

• DMPK dimerization, mediated by the H domain, significantly influences its catalytic efficiency and substrate binding.
• These dimerization-dependent functional changes are likely important for DMPK's role in myotonic dystrophy.
• The findings suggest similar dimerization mechanisms may be relevant for other protein kinases with coiled-coil domains.