The protein phosphatases involved in cellular regulation. 1. Classification and substrate specificities

Area of Science:

• Biochemistry
• Enzymology
• Metabolic Regulation

Background:

• Protein phosphatases are crucial for regulating intermediary metabolism.
• Four main enzymes, divided into type-1 and type-2, are responsible for these activities.
• Type-1 phosphatases are inhibited by inhibitor-1 and -2, while type-2 are insensitive.

Purpose of the Study:

• To investigate the substrate specificities of four key protein phosphatase enzymes: protein phosphatase-1 (type-1) and protein phosphatases 2A, 2B, and 2C (type-2).
• To understand how these enzymes regulate major metabolic pathways.
• To explore the determinants of protein phosphatase specificity.

Main Methods:

• Phosphorylation of 13 different substrate proteins using eight different protein kinases.
• Assay of dephosphorylation activities of protein phosphatases 1, 2A, 2B, and 2C on various substrates.
• Analysis of substrate specificities and enzyme inhibition patterns.

Main Results:

• Protein phosphatase-1 and 2A exhibit broad substrate specificities across diverse metabolic pathways.
• Protein phosphatase-2A shows higher activity on myosin light chain and ATP-citrate lyase.
• Protein phosphatase-2C has distinct low activities on phosphorylase and histone H1, but high activity on HMG-CoA reductase.
• Protein phosphatase-2B is highly specific, acting primarily on phosphorylase kinase alpha-subunit, inhibitor-1, and myosin light chains, and is inhibited by trifluoperazine.

Conclusions:

• Protein phosphatase-1 and 2A are major regulators of intermediary metabolism due to their broad substrate specificities.
• Protein phosphatase-2B and 2C have more specialized roles in metabolic control.
• Primary amino acid sequence near phosphorylation sites does not solely determine protein phosphatase specificity.