Insulin-receptor phosphotyrosyl-protein phosphatases

Area of Science:

• Biochemistry
• Molecular Biology
• Cell Signaling

Background:

• Calmodulin-dependent protein phosphatase (CaM-PPase) is implicated in phosphotyrosyl-protein phosphatase activity.
• The role of CaM-PPase in dephosphorylating insulin and epidermal growth factor (EGF) receptors requires further investigation.

Purpose of the Study:

• To compare the dephosphorylation activity of CaM-PPase on insulin receptor versus EGF receptor.
• To characterize native phosphotyrosyl-protein phosphatase activity in cell extracts.
• To determine the contribution of CaM-PPase to overall phosphotyrosyl-protein phosphatase activity in cellular fractions.

Main Methods:

• Enzymatic assays using purified CaM-PPase and 32P-labeled insulin and EGF receptors.
• Characterization of native phosphatase activity in particulate and soluble fractions of rat liver, heart, and brain extracts.
• Metal ion dependence studies, specifically using Ni2+, to differentiate CaM-PPase activity.

Main Results:

• Purified CaM-PPase dephosphorylated both insulin and EGF receptors, with higher activity against EGF receptor.
• Native phosphotyrosyl-protein phosphatase activity against both receptors was predominantly found in the particulate fraction (75%) of cell extracts.
• Ni2+ strongly inhibited both particulate and soluble phosphotyrosyl-protein phosphatase activity, indicating CaM-PPase is not the major phosphatase for these receptors in cell extracts.

Conclusions:

• Calmodulin-dependent protein phosphatase can dephosphorylate both insulin and EGF receptors, but at different rates.
• The majority of phosphotyrosyl-protein phosphatase activity against insulin and EGF receptors in rat tissues resides in the particulate fraction.
• Phosphotyrosyl-protein phosphatases other than CaM-dependent ones are the principal enzymes responsible for dephosphorylating insulin and EGF receptors in cell extracts.