Ligand-triggered conformational perturbations elicit changes at the single cysteinyl residue of spinach calmodulin.
Following application of stoichiometric amounts of Ca2+ or specific partner peptides to spinach calmodulin, dynamic changes in the nanosecond range could be monitored at a strategically anchored fluorescence or spin probe. For these studies the single cysteinyl residue 26 of spinach calmodulin was labelled with a thiol-specific proxyl (i.e. 2,2,5,5-tetramethyl-1-pyrrolidinyl-oxyl) spin probe or with a bimane fluorescence probe. With Ca2+ and a specific ligand (mastoparan) present, fluorescence studies (anisotropy, lifetime) indicated that the rotational motion of the protein complex becomes slower relative to the motion of calmodulin in the absence of the specific ligand. The probe's attachment site 26 appears to reside in a fairly polar microenvironment as reported by a series of proxyl spin probes varying in label length. The rotational correlation time of the shortest spin probe markedly changed upon binding of a specific peptide to a calmodulin region distant from that of the monitoring spin probe. We interpret these observations as indicating that ligand-triggered conformational perturbations are eliciting specific responses at the cysteinyl residue 26 of spinach calmodulin.
Yuan
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SX
Department of Microbiology, Michigan State University, East Lansing 48824-1311.
Haug
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A
D001514:Bee Venoms / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism*; D002147:Calmodulin; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D003545:Cysteine; D004578:Electron Spin Resonance Spectroscopy; D005453:Fluorescence; D036341:Intercellular Signaling Peptides and Proteins; D008024:Ligands / Q000378:metabolism; D010455:Peptides; D010944:Plants; D011487:Protein Conformation; D014862:Wasp Venoms / Q000378:metabolism*
D001514:Bee Venoms; D002147:Calmodulin; D036341:Intercellular Signaling Peptides and Proteins; D008024:Ligands; D010455:Peptides; D014862:Wasp Venoms; C037500:mastoparan; D003545:Cysteine; D002118:Calcium