Type III metacaspases: calcium-dependent activity proposes new function for the p10 domain

Area of Science:

• Biochemistry
• Molecular Biology
• Algal Biology

Background:

• Metacaspases are caspase homologues found in bacteria, algae, and plants.
• Type I and II metacaspases are in plants, while type III metacaspases are unique to algae with secondary endosymbiosis.
• The function and characteristics of type III metacaspases remain largely uncharacterized.

Purpose of the Study:

• To analyze the expression of caspase homologues in the cryptophyte Guillardia theta.
• To biochemically characterize the type III metacaspase GtMC2 from G. theta.
• To investigate the functional relationship between type III metacaspases and other metacaspase types.

Main Methods:

• In vivo expression analysis of 13 caspase homologues in Guillardia theta.
• In vitro biochemical characterization of the type III metacaspase GtMC2.
• Identification and analysis of calcium-binding sites and their impact on enzyme activity.

Main Results:

• Type III metacaspase GtMC2 functions as an endopeptidase with a preference for basic amino acids at the P1 position.
• Full catalytic efficiency of GtMC2 requires specific N-terminal proteolytic cleavage and calcium ions (low millimolar concentrations).
• Two calcium-binding sites were identified in GtMC2, with dissociation constants in low and high micromolar ranges, suggesting a low-affinity site in the p10 domain.

Conclusions:

• Type III metacaspases show high functional relatedness to type I metacaspases.
• The conserved N-terminal region in the p10 domain, present in type I/II/III metacaspases, is crucial for calcium-dependent activity.
• This study provides significant insights into the structure, function, and evolutionary significance of type III metacaspases in algae.