Methionine to glutamine substitutions in the C-terminal domain of calmodulin impair the activation of three protein kinases.
The 9 methionine residues of vertebrate calmodulin (CaM) were individually changed to glutamine residues in order to investigate their roles in enzyme binding and activation. The mutant proteins showed three classes of effect on the activation of smooth muscle myosin light chain kinase, CaM-dependent protein kinase IIalpha, and CaM-dependent protein kinase IV. First, some mutations had no appreciable effect on the ability of CaM to activate the three protein kinases. Included in this category were glutamine substitutions at residues 36 and 51 in the N-terminal domain, at residue 76 in the domain linker sequence, and at residues 144 and 145 in the C-terminal domain. Second, glutamine substitutions in the N-terminal domain of CaM, particularly those at positions 71 and 72, lowered the maximal activity of smooth muscle myosin light chain kinase while having no effect on the other two enzymes. Finally the affinity of CaM for all three enzymes was lowered by glutamine mutations at the neighboring methionines 109 and 124, located on a solvent-accessible surface of the C-terminal domain of Ca2+/CaM. This last result provides the first demonstration of the involvement of the same hydrophobic groups in the high affinity binding of CaM to three different enzymes.
Chin
D
D
Department of Pharmacology, Duke University Medical Center, Durham, North Carolina 27710, USA.
Means
A R
AR
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002118:Calcium / Q000502:physiology; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D054734:Calcium-Calmodulin-Dependent Protein Kinase Type 4; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000737:chemistry / Q000378:metabolism*; D002147:Calmodulin / Q000737:chemistry* / Q000378:metabolism; D004789:Enzyme Activation; D005973:Glutamine / Q000737:chemistry; D007700:Kinetics; D008715:Methionine / Q000737:chemistry; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009219:Myosin-Light-Chain Kinase / Q000737:chemistry / Q000378:metabolism*; D010455:Peptides / Q000737:chemistry; D010766:Phosphorylation; D011485:Protein Binding; D013329:Structure-Activity Relationship
D002147:Calmodulin; D010455:Peptides; D005973:Glutamine; D008715:Methionine; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D054734:Calcium-Calmodulin-Dependent Protein Kinase Type 4; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D009219:Myosin-Light-Chain Kinase; D002118:Calcium