251. Fourier transform infrared spectroscopic study on the binding of Mg2+ to a mutant Akazara scallop troponin C (E142Q)
- Author
-
Takao Ojima, Koji Nagata, Fumiaki Yumoto, Masaru Tanokura, Hiroyuki Kagi, Kiyoyoshi Nishita, and Masayuki Nara
- Subjects
Biochemical Phenomena ,Biophysics ,Analytical chemistry ,Infrared spectroscopy ,Biochemistry ,Biomaterials ,Troponin C ,Troponin complex ,Spectroscopy, Fourier Transform Infrared ,medicine ,Animals ,Magnesium ,Fourier transform infrared spectroscopy ,Binding site ,Chemistry ,Circular Dichroism ,Muscles ,Organic Chemistry ,Wild type ,Skeletal muscle ,General Medicine ,musculoskeletal system ,Crystallography ,medicine.anatomical_structure ,Spectrometry, Fluorescence ,Mollusca ,Scallop ,Mutation ,Protein Binding - Abstract
Troponin C (TnC) is the Ca 2 + -binding regulatory protein of the troponin complex in muscle tissue. Vertebrate fast skeletal muscle TnCs bind four Ca 2 + , while Akazara scallop (Chlamys nipponensis akazara) striated adductor muscle TnC hinds only one Ca 2 + at Site IV, because all the other EF-hand motifs are short of critical residues for the coordination of Ca 2 + . Fourier transform infrared (FTIR) spectroscopy was applied to study coordination structure of Mg 2 + bound in a mutant Akazara scallop TnC (E142Q) in D 2 O solution. The result showed that the side-chain COO groups of Asp 131 and Asp 133 in the Ca 2 + -binding site of E142Q bind to Mg 2 + in the pseudo-bridging mode. Mg 2 + titration experiments for E142Q and the wild-type of Akazara scallop TnC were performed by monitoring the band at about 1600 cm - 1 , which is due to the pseudo-bridging Asp COO groups. As a result, the binding constants of them for Mg 2 + were the same value (about 6 mM). Therefore, it was concluded that the side-chain COO group of Glu 142 of the wild type has no relation to the Mg 2 + ligation. The effect of Mg 2 + boding in E142Q was also investigated by CD and fluorescence spectroscopy. The on-off methanism of the activation of Akazara scallop TnC is discussed on tile basis of the coordination structures of Mg 2 + as well as Ca 2 + .
- Published
- 2004