Abstract
In an attempt to draw a correlation between calcium carbonate (CaCO3) precipitation and biomacromolecules such as extracellular polymeric substances and enzyme activity in biomineralizing microbe, this report aims to elucidate the ureolytic and ammonification route in Paenibacillus alkaliterrae to explore the possible role of organic biomolecule(s) present on cell surface in mediating nucleation and crystallization of biogenic CaCO3. After 168 h of biomineralization in ureolysis and ammonification, 2.2 g/l and 0.87 g/l of CaCO3 precipitates were obtained, respectively. The highest carbonic anhydrase activity (31.8 µmoles/min/ml) was evidenced in ammonification as opposed to ureolysis (24.8 µmoles/min/ml). Highest urease activity reached up to 9.26 µmoles/min/ml in ureolytic pathway. Extracellular polymeric substances such as polysaccharides and proteins were found to have a vital role not only in the nucleation and crystal growth but also in addition direct polymorphic fate of CaCO3 nanoparticles. EPS production was higher during ammonification (3.1 mg/ml) than in ureolysis (0.72 mg/ml). CaCO3 nanoparticle–associated proteins were found to be 0.82 mg/ml in ureolysis and 0.56 mg/ml in ammonification. After 30 days of biomineralization, all the polymorphic forms stabilized to calcite in ureolysis but in ammonification vaterite predominated. In our study, we showed that organic template–mediated prokaryotic biomineralization follows the non-classical nucleation and varying proportions of these organic components causes selective polymorphism of CaCO3 nanoparticles. Overall, the findings are expected to further the fundamental understanding of enzymes, EPS-driven non-classical nucleation of CaCO3, and we foresee the design of fit-for-purpose futuristic biominerals arising from such renewed understanding of biomineralization.
Key points
• Organic-inorganic interface of cell surface promote crystallization of biominerals
• Carbohydrate and proteins in the interface results selective polymorphism of CaCO3
• Calcite stabilized at 30 days in ureolysis, vaterite-calcite mix in ammonification
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Acknowledgements
The authors thank the Department of Biotechnology and Central Research Facility (CRF) of IIT Kharagpur for providing necessary research facilities. BioRender.com is duly acknowledged for creation of images. The authors are thankful to Prof. Abhijit Mukherjee, John Curtin Distinguished Professor, Structural Engineering, School of Civil and Mechanical Engineering and Dr. Navdeep Dhami, Senior Lecturer, School of Molecular and Life Sciences, Curtin University, Perth, Australia for their insightful scientific inputs during their visit to IIT Kharagpur.
Funding
The authors gratefully acknowledge the DBT-JRF fellowship (Fellow No. DBT/2018/IIT-KH/1012) New Delhi, India, to Ankita Debnath and Scheme of Young Scientists and Technologists (SYST) (File no.: SP/YO/530/2018, 08/10/2018), Department of Science and Technology (DST), Government of India to Chinmay Hazra for financial support. The authors also acknowledge financial assistance from 'Scheme for Promotion of Academic and Research Collaboration' [SPARC, MHRD, New Delhi] (SPARC/2018-2019/P409/SL,Dt.11-07-2019).
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RS: conceptualization, validation, writing—review and editing draft, resources, and supervision; AD: experiments and analysis, methodology, validation, and writing—original draft; CH: validation and writing—review and editing draft.
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Debnath, A., Hazra, C. & Sen, R. Insight into biomolecular interaction–based non-classical crystallization of bacterial biocement. Appl Microbiol Biotechnol 107, 6683–6701 (2023). https://doi.org/10.1007/s00253-023-12736-5
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DOI: https://doi.org/10.1007/s00253-023-12736-5