Subject(s)
Metals , Chemistry, Inorganic , History, 20th Century , History, 21st Century , United KingdomSubject(s)
Calcineurin Inhibitors , Intracellular Signaling Peptides and Proteins/metabolism , Isoenzymes/metabolism , Multigene Family , Muscle Proteins/metabolism , Animals , Calcineurin/metabolism , DNA-Binding Proteins , Down Syndrome , Humans , Intracellular Signaling Peptides and Proteins/genetics , Isoenzymes/genetics , Muscle Proteins/geneticsABSTRACT
Previous studies have claimed that there is a failure of a mutant form of superoxide dismutase (mSOD) to protect the protein phosphatase, calcineurin (CN), against inactivation in the pathogenesis of amyotrophic lateral sclerosis (ALS), as determined in a murine model of ALS resulting from overexpression of mSOD (G93A). In contrast to previous studies, we find that mice overexpressing G93A mSOD have no statistically significant differences in the expression, or activity, of CN. However, CN from G93A mSOD overexpressing mice is significantly more protected against inactivation than non-transgenic mice that do not overexpress SOD. This reduced inactivation of CN is a consequence of increased expression of G93A mSOD. Thus, like wild-type SOD, G93A mSOD protects CN against inactivation.
Subject(s)
Amyotrophic Lateral Sclerosis/metabolism , Calcineurin/metabolism , Point Mutation , Superoxide Dismutase/genetics , Superoxide Dismutase/metabolism , Amyotrophic Lateral Sclerosis/genetics , Amyotrophic Lateral Sclerosis/physiopathology , Animals , Humans , Mice , Mice, TransgenicSubject(s)
Calcineurin/metabolism , Oxidative Stress/physiology , Amino Acid Sequence , Animals , Brain/enzymology , Calcineurin Inhibitors , Calcium/metabolism , Calmodulin/antagonists & inhibitors , Calmodulin/metabolism , Cattle , Chromatography, Ion Exchange/methods , Erythrocytes/enzymology , Indicators and Reagents , Iron/pharmacology , Kinetics , Molecular Sequence Data , Peptide Fragments/chemistry , Peptide Fragments/pharmacology , Phosphorylation , Substrate Specificity , Superoxide Dismutase/blood , Zinc/pharmacologyABSTRACT
Edema factor (EF), a toxin from Bacillus anthracis (anthrax), possesses adenylyl cyclase activity and requires the ubiquitous Ca2+-sensor calmodulin (CaM) for activity. CaM can exist in three major structural states: an apo state with no Ca2+ bound, a two Ca2+ state with its C-terminal domain Ca2+-loaded, and a four Ca2+ state in which the lower Ca2+ affinity N-terminal domain is also ligated. Here, the interaction of EF with the three Ca2+ states of CaM has been examined by NMR spectroscopy and changes in the Ca2+ affinity of CaM in the presence of EF have been determined by flow dialysis. Backbone chemical shift perturbations of CaM show that EF interacts weakly with the N-terminal domain of apoCaM. The C-terminal CaM domain only engages in the interaction upon Ca2+ ligation, rendering the overall interaction much tighter. In the presence of EF, the C-terminal domain binds Ca2+ with higher affinity, but loses binding cooperativity, whereas the N-terminal domain exhibits strongly reduced Ca2+ affinity. As judged by chemical shift differences, the N-terminal CaM domain remains bound to EF upon subsequent Ca2+ ligation. This Ca2+ dependence of the EF-CaM interaction differs from that observed for most other CaM targets, which normally interact only with the Ca2+-bound CaM domains and become active following the transition to the four Ca2+ state.
