Objectives: To overcome the limited production capability of shell matrix proteins and efficiently conduct in vitro CaCO3 biomineralization studies, a putative recombinant shell matrix protein was prepared and simply characterized.

Results: A glycine-rich protein (named GRP_BA) was found as a putative shell matrix protein (NCBI reference sequence; BAA20465), and the protein was genetically redesigned for the recombinant production in Escherichia coli. The recombinant protein was successfully obtained in the bacterial expression system with the final yield of approximately 30 mg L-1 and the purity of above 95% in a 400-mL lab scale flask culture. This protein was able to efficiently form a complex with calcium ions (Ca2+), and Ca2+-induced agglomeration was observed like other calcification-related proteins. Spherulitic calcite micro-particles of 20-30 µm diameter with rosette- and sphere-like structures were synthesized in the presence of the recombinant shell protein, which could be formed by stacking and/or aggregation of calcite nanograins and the bound protein.

Conclusions: Recombinant production of the shell matrix protein could overcome potential difficulties associated with limited amount of protein available for biomineralization studies and provide opportunities to fabricate notable biominerals in practical aspects.