Contributions of myristoylation to calcineurin structure/function.
Calcineurin is a serine/threonine protein phosphatase composed of a catalytic subunit, calcineurin A (58 kDa), and a NH2-terminal myristoylated regulatory subunit, calcineurin B (19 kDa). In order to study the effect of myristoylation on calcineurin structure/function, a dual plasmid transfection system was used to generate myristoylated and nonmyristoylated calcineurin B. Both metabolic labeling of calcineurin B with radiolabeled myristic acid and electrospray mass spectral analysis confirmed that myristic acid was covalently and stoichiometrically linked to calcineurin B. Myristoyl and non-myristoyl calcineurin B were reconstituted with recombinant calcineurin A to form native-like heterodimers, and the properties of the two calcineurin forms were examined. Myristoylation had no effect on enzymatic activity, calcineurin-immunosuppressant/immunophilin interactions, or Ca2+ binding. Surprisingly, myristoylation also had no effect on calcineurin heterodimer association with phospholipid monolayers. Fatty acylation, however, significantly influenced the thermal stability of calcineurin, with an approximate 10 degrees C increase in t1/2 observed for myristoyl calcineurin when compared to the non-myristoyl form. Myristoylation of calcineurin B therefore appears to provide structural stability to the calcineurin heterodimer.
Section of Hematology Research and the Department of Biochemistry and Molecular Biology, Mayo Clinic and Mayo Foundation, Rochester, Minnesota 55905, USA.
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D000591:Amino Acid Isomerases / Q000494:pharmacology; D000818:Animals; D019703:Calcineurin; D002148:Calmodulin-Binding Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism*; D002352:Carrier Proteins / Q000494:pharmacology; D021983:Cyclophilins; D016572:Cyclosporine / Q000494:pharmacology; D004795:Enzyme Stability; D006358:Hot Temperature; D019814:Myristic Acid; D009227:Myristic Acids / Q000378:metabolism*; D019696:Peptidylprolyl Isomerase; D010743:Phospholipids / Q000378:metabolism; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism*; D051381:Rats; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism; D013329:Structure-Activity Relationship
D002148:Calmodulin-Binding Proteins; D002352:Carrier Proteins; D009227:Myristic Acids; D010743:Phospholipids; D011994:Recombinant Proteins; D019814:Myristic Acid; C081786:cyclophilin B; D016572:Cyclosporine; D019703:Calcineurin; D010749:Phosphoprotein Phosphatases; D000591:Amino Acid Isomerases; D021983:Cyclophilins; D019696:Peptidylprolyl Isomerase