Ca2+ differentially regulates conventional protein kinase Cs' membrane interaction and activation

Area of Science:

• Biochemistry
• Molecular Biology
• Enzymology

Background:

• Conventional protein kinase Cs (PKCs) are crucial signaling enzymes regulated by calcium ions (Ca2+).
• Understanding the precise mechanisms of Ca2+ regulation is vital for deciphering cellular signaling pathways.

Purpose of the Study:

• To investigate how Ca2+ affects the membrane binding, catalytic function, and conformation of conventional PKCs.
• To elucidate the role of specific protein kinase C isozymes and their structural domains in Ca2+ regulation.

Main Methods:

• Enzyme kinetics assays to determine Ca2+ concentrations for half-maximal membrane binding and activation.
• Protease-mediated conformational probing to analyze structural changes induced by Ca2+.
• Comparative analysis of protein kinase C betaI and betaII isozymes.

Main Results:

• Lower Ca2+ concentrations are required for membrane binding than for catalytic activation.
• Protein kinase C betaII exhibits distinct Ca2+ binding and activation kinetics compared to betaI, with carboxyl-terminal residues influencing Ca2+ regulation.
• Ca2+-dependent membrane binding and activation involve separate conformational changes across multiple protein domains, including hinge motion and terminus exposure/masking.

Conclusions:

• Isozyme-specific structural features, including those in the carboxyl terminus, determine protein kinase C's Ca2+-dependent affinity for membranes.
• Distinct structural rearrangements govern Ca2+ binding and catalytic activity, highlighting complex interdomain communication within PKCs.