Reassessing enzyme kinetics: Considering protease-as-substrate interactions in proteolytic networks

Area of Science:

• Biochemistry
• Enzymology
• Systems Biology

Background:

• Enzymes, as biological catalysts, are typically assumed to be inert and unaltered during reactions.
• Proteases, a specific enzyme class, hydrolyze other proteins and can act as substrates for other proteases.
• Multienzyme systems, especially those involving proteases, exhibit complex dynamics where enzyme self-degradation impacts overall kinetics.

Purpose of the Study:

• To challenge the assumption of enzyme inertness in protease systems.
• To investigate the impact of protease-on-protease interactions on kinetic predictions.
• To develop a more accurate model for proteolytic network dynamics.

Main Methods:

• Investigated protease-on-protease inactivating hydrolysis and "cathepsin cannibalism" dynamics.
• Utilized computational models to explore proteolytic network interactions, including cooperative/competitive degradation and substrate cleavage.
• Incorporated additional reaction terms such as autodigestion, inactivation, and "distraction" reactions (inhibitory effects of inactivated proteases).

Main Results:

• Protease self- and cross-degradation significantly alters predicted protease concentrations and substrate degradation rates.
• "Distraction reactions," where inactivated proteases inhibit active ones, were identified as a key network node.
• The inclusion of these complex dynamics improved the accuracy of proteolytic network predictions.

Conclusions:

• Enzyme loss through proteolytic degradation must be considered in multiprotease systems, altering standard enzymatic kinetic models.
• Accurate modeling of protease networks requires incorporating self- and cross-inactivation mechanisms.
• An online platform was developed to model these complex cathepsin network dynamics, aiding research and pharmaceutical dosing strategy development.