Incorporation of fluorescent molecules and proteins into calcium oxalate monohydrate single

A central question to understanding biomineralization is determining how bi molecules are integrated within inorganic host lattices, thereby changing m aterial properties yet retaining single crystal structure of the biomineral . We have addressed guest incorporation within single biomineral crystals b y investigating face specificity, anisotropy and the role of charges in flu orescent calcium oxalate monohydrate (COM) crystals nucleated from solution s containing eosin Y, fluoresceins and rhodamines. Additionally, we have ex amined the specificity of incorporation of Protein G wild-type (G-wt) and i ts mutant (G-Delta6, in which four aspartic acids and two glutamic acids ha ve been replaced by the corresponding asparagine or glutamine), where G-wt promotes and G-Delta6 inhibits COM crystal growth. We found that (1) the ne gatively charged fluorophores, as well as the fluorophore-labeled proteins, G-wt and G-Delta6, were successfully incorporated during growth into the s ame {101} growth sectors in preference to all others, (2) the positively ch arged TRITC (tetramethyl rhodamine isothiocyanate) was not incorporated as free fluorophore, but it became incorporated if conjugated to G-wt and G-De lta6; (3) once the fluorophores are incorporated, the polarization measurem ents of adsorption and emission were similar irrespectively whether taken f rom COM containing the free fluorophores, or the protein conjugates. The an isotropy was similar for rhodamines and fluoresceins. (C) 2001 Elsevier Sci ence B.V. All rights reserved.