Cation binding properties of calretinin, an EF-hand calcium-binding protein.
Calretinin (CR) is a neuronal EF-hand protein previously characterized as a calcium (micromolar affinity) binding protein. CR-containing neurons are spared in some neurodegenerative diseases, although it is as yet unconfirmed how CR plays an active role in this protection. Higher levels of some metal cations (e.g. copper and zinc) are associated with these diseases. At the same time, metals such as terbium (NMR and fluorescence) cadmium (NMR) and manganese (EPR) serve as useful calcium analogues in the study of EF-hand proteins. We survey the binding of the above-mentioned metal cations that might affect the structure and function of CR. Competitive 45Ca2+-overlay, competitive terbium fluorescence and intrinsic tryptophan fluorescence are used to detect the binding of metal cations to CR. Terbium and copper (half-maximal effect of 15 microM) bind to CR. Terbium has a similar or greater affinity for the calcium-binding sites of CR than calcium. Copper quenches the fluorescence of terbium-bound CR, and CR tryptophan residues and competes weakly for 45Ca2+-binding sites. Cadmium, magnesium, manganese and zinc bind less strongly (half-maximal effects above 0.1 mM). Therefore, only terbium appears to be a suitable analytical calcium analogue in further studies of CR. The principal conclusion of this work is that copper, in addition to calcium, might be a factor in the function of CR and a link between CR and neurodegenerative diseases.
Groves
P
P
Laboratory of Calcium-Binding Proteins, Nencki Institute of Experimental Biology, Warszawa, Poland. patrick@nencki.gov.pl
Palczewska
M
M
D000818:Animals; D001667:Binding, Competitive; D002104:Cadmium / Q000737:chemistry; D064032:Calbindin 2; D002118:Calcium / Q000378:metabolism; D002412:Cations; D004305:Dose-Response Relationship, Drug; D004578:Electron Spin Resonance Spectroscopy; D006863:Hydrogen-Ion Concentration; D009682:Magnetic Resonance Spectroscopy; D008345:Manganese / Q000737:chemistry; D019636:Neurodegenerative Diseases / Q000378:metabolism; D011485:Protein Binding; D051381:Rats; D064030:S100 Calcium Binding Protein G / Q000737:chemistry* / Q000378:metabolism*; D013050:Spectrometry, Fluorescence; D013725:Terbium / Q000737:chemistry*; D014364:Tryptophan / Q000737:chemistry
C576512:Calb2 protein, rat; D064032:Calbindin 2; D002412:Cations; D064030:S100 Calcium Binding Protein G; D002104:Cadmium; D013725:Terbium; D008345:Manganese; D014364:Tryptophan; D002118:Calcium