Effects of calcium and calcium analogs on calmodulin: a Fourier transform infrared and electron spin resonance investigation.
Fourier transform infrared (FTIR) and electron spin resonance (ESR) spectroscopies have been used to monitor changes in the conformation of calmodulin induced by Ca2+ and Ca2+ analogs. Using FTIR spectroscopy we observe that Ca2+: (i) favors the alpha-helical conformation and decreases the flexibility of the molecule; (ii) multiplies the intramolecular hydrogen bonds (the ratio of freely vibrating NH/hydrogen bound NH groups decreases); (iii) induces changes in the C-terminal tyrosine environment; and (iv) increases compactness of the molecule (less NH groups in the peptide bonds can be deuterated). As proved by ESR, Ca2+ binding induces exposure of hydrophobic domains allowing binding of a spin-labelled phenothiazine on calmodulin. When the experiments are performed in the presence of increasing amounts of Ca2+, both ESR and FTIR provide evidence that major conformational changes result after the filling of only two Ca2+-binding sites. But achievement of the spectroscopical changes is only observed when the four binding sites are filled (Ca2+/calmodulin = 4). The effects of analogs are monitored with the same spectroscopical parameters. Zn+ does not induce structural modifications of calmodulin but all other analogs studied mimic the calcium effects to some extent. As regards the amplitude of the spectroscopical effects, analogs rank in the following order: Ca2+ greater than Cd2+ greater than Tb3+ = Eu3+ greater than Gd3+ greater than La3+ greater than Zn2+ = cation depleted. Except for Zn2+, ranking for their activating potency of MLCK, the analogs can be arranged in a similar order.
Rainteau
D
D
Département de Biochimie, UER Biomédicale des Saints-Pères, Paris, France.
Wolf
C
C
Lavialle
F
F
D000818:Animals; D002104:Cadmium / Q000494:pharmacology; D002118:Calcium / Q000494:pharmacology*; D002147:Calmodulin / Q000494:pharmacology; D002412:Cations; D004578:Electron Spin Resonance Spectroscopy; D004789:Enzyme Activation / Q000187:drug effects; D005063:Europium / Q000494:pharmacology; D005682:Gadolinium / Q000494:pharmacology; D008297:Male; D009219:Myosin-Light-Chain Kinase / Q000378:metabolism; D011487:Protein Conformation / Q000187:drug effects; D011817:Rabbits; D012756:Sheep; D013055:Spectrophotometry, Infrared; D013725:Terbium / Q000494:pharmacology; D013737:Testis / Q000032:analysis; D015032:Zinc / Q000494:pharmacology
D002147:Calmodulin; D002412:Cations; D002104:Cadmium; D013725:Terbium; D005063:Europium; D005682:Gadolinium; D009219:Myosin-Light-Chain Kinase; D015032:Zinc; D002118:Calcium