A novel urease gene structure of Sporosarcina pasteurii with double operons.

Microbially induced calcium carbonate precipitation (MICP) had emerged as an important biomineralization process with wide-ranging applications in construction, environmental remediation, and space exploration. Sporosarcina pasteurii (S. pasteurii) was a key bacterium in MICP due to its efficient urease activity, yet the regulation of its urease genes remains poorly understood, limiting its practical applications. This study aimed to elucidate the structure and expression regulation mechanism of urease genes in S. pasteurii to enhance its mineralization potential. We compared the growth and urease gene expression of S. pasteurii under three different culture conditions using transcriptome sequencing. Operon, Transcription Start Site (TSS) and Transcription Termination Site (TTS) were predicted based on the distribution of reads on the genome using Rockhopper online analysis software. The 700 bp sequence upstream of the TTS was extracted and promoter prediction was performed by Time-Delay Neural Network (TDNN) method. Finally, we verified the prediction results by RT-PCR. Our results revealed, for the first time, a double operon structure of S. pasteurii urease, with operon 1 containing ureA, ureB, ureC, ureE, and ureF genes, and operon 2 containing ureG and ureD genes. This discovery provides crucial insights into the regulation of urease expression in S. pasteurii, paving the way for more efficient and controllable mineralization applications. The findings of this study not only advanced our understanding of urease gene regulation but also opened new avenues for optimizing S. pasteurii-based biomineralization technologies.