The Mechanism Of Amorphous Calcium Carbonate Formation And Transformation Controlled By ACCBP

Amorphous calcium carbonate (ACC) plays an important role in biomineralizationprocess for its function as a precursor for calcium carbonate biominerals. Themechanisms for ACC regulation is one of the most interesting questions in the field ofbiomineralization. In this thesis, we studied the function of amorphous calciumcarbonate-binding (ACCBP) during ACC formation and transformation, and themechanisms for it.Biochemical experiments coupled with bioinformatics approaches were usded toexplore the mechanisms of ACC formation controlled by ACCBP. Size-exclusionchromatography, chemical cross-linking experiments and negative staining electronmicroscopy reveal that ACCBP is a decamer composed of two adjacent pentamers.Sequence analyses and fluorescence quenching results indicate that ACCBP containstwo Ca2+-binding sites. The results of in vitro crystallization experiments suggest thatone Ca2+-binding site is critical for ACC formation and the other site affects the ACCinduction efficiency. As homologous proteins of ACCBP are all pentamers and ACCBPitself is a decamer comprosed of two pentamers, a homology modeling was conductedwith the structure of a pentermeric protein, α-7nicotinic receptor. Homology modelingdemonstrates that the Ca2+-binding sites of pentameric ACCBP are arranged in a five-fold symmetry, which is the structural basis for ACC formation. To the best of ourknowledge, this is the first report on the structural basis for protein-induced ACCformation and it will significantly improve our understanding of the amorphousprecursor pathway.Another key question in the amorphous precursor pathway is the polymorphdetermination which is delicately controlled by biomacromolecules, especially proteins.However, the mechanism of protein-mediated polymorph determination is still unclear.In this thesis, in vitro crystallization experiments and ACC transformation experimentsshow that ACCBP induces aragonite via ACC precursor in solution with low Mg/Ca atlow temperature. SEM images show that the ACC precursor in the ACC transformationexperiment is nanograins. XPS, ICP, Raman, and FTIR analyses on the ACC nanograins show that ACCBP increases Mg/Ca molar ratio on the surface of ACC, butdoes not regulate the bulk Mg/Ca ratio or the short-range order structure. It is proposedthat the polymorph switching is controlled by increasing the surficial Mg/Ca of transientACC nanograins, and it may be the mechanism for polymorph selection in vivo. Theseresults would contribute to the understanding of biomineralization, particularly howprotein cooperate with Mg2+on the transformation of amorphous calcium carbonate.In summary, the functions of ACCBP in the biomineralization process werestudied and the mechanisms for ACC formation and polymorph determination in theamorphous precursor pathway were discussed in this thesis.