Circularly polarized luminescence from terbium(III) as a probe of metal ion binding in calcium-binding proteins.
A number of different experimental techniques have been used to probe the details of structural changes on the binding of Ca(II) to the large number of known calcium-binding proteins. The use of luminescent lanthanide(III) ions, especially terbium(III) and europium(III), as substitutional replacement for calcium(II), has led to a number of useful experiments from which important details concerning the metal ion coordination sites have been obtained. This work is concerned with the measurement of the circularly polarized luminescence (CPL) from the 5D4----7F5 transition of Tb(III) bound to the calcium binding sites of bovine trypsin, bovine brain calmodulin, and frog muscle parvalbumin. It is demonstrated that it is possible to make these polarization measurements from very dilute solutions (less than 20 microM) and monitor structural changes as equivalents of Tb(III) are added. It is shown that the two proteins that belong to the class of "EF-hand" structures (calmodulin and parvalbumin) possess quite similar CPL line shapes, whereas Tb(III) bound to trypsin has a much different band structure. CPL results following competitive and consecutive binding of Ca(II) and Tb(III) bound to calmodulin are also reported and yield information concerning known differences between the sequence of binding of these two species.
Coruh
N
N
Department of Chemistry, University of Missouri, St. Louis 63121.
Riehl
J P
JP
D001665:Binding Sites; D001667:Binding, Competitive; D001923:Brain Chemistry; D002118:Calcium / Q000378:metabolism; D002135:Calcium-Binding Proteins / Q000737:chemistry* / Q000378:metabolism; D002147:Calmodulin / Q000737:chemistry / Q000378:metabolism; D008163:Luminescent Measurements; D009132:Muscles / Q000737:chemistry; D010320:Parvalbumins / Q000737:chemistry / Q000378:metabolism; D011487:Protein Conformation; D013057:Spectrum Analysis; D013725:Terbium / Q000378:metabolism*; D014357:Trypsin / Q000737:chemistry / Q000378:metabolism
D002135:Calcium-Binding Proteins; D002147:Calmodulin; D010320:Parvalbumins; D013725:Terbium; D014357:Trypsin; D002118:Calcium