Part:BBa_K5178006

Coding sequence for pancreatic procolipase enzyme with a signal peptide and histidine purification tag. This part is under the control of the constitutive pVEG promoter (Part:BBa_K823003). The part sequence is codon optimized for expression in B. subtilis. Addition of a signal peptide allows the enzyme to be more efficiently secreted by transformed B. subtilis. A HA purification tag was added for easy enzyme detection and quantification during experimental assays. This part also includes coding sequences for RiboJ—a ribozyme that increases access to the adjacent RBS, a generic RBS: MF001, and a generic terminator: B0015.

Coding sequences were gathered from the following sources.

CLPS: van Tilbeurgh, H., Sarda, L., Verger, R. et al. Structure of the pancreatic lipase–procolipase complex. Nature 359, 159–162 (1992). https://doi.org/10.1038/359159a0

PVEG: Part:BBa_K823003

RiboJ: Lou, C., Stanton, B., Chen, Y. J., Munsky, B., & Voigt, C. A. (2012). Ribozyme-based insulator parts buffer synthetic circuits from genetic context. Nature biotechnology, 30(11), 1137–1142. https://doi.org/10.1038/nbt.2401

MF001: Castillo-Hair, S. M., Fujita, M., Igoshin, O. A., & Tabor, J. J. (2019). An Engineered B. subtilis Inducible Promoter System with over 10 000-Fold Dynamic Range. ACS synthetic biology, 8(7), 1673–1678. https://doi.org/10.1021/acssynbio.8b00469

B0015: Castillo-Hair, S. M., Fujita, M., Igoshin, O. A., & Tabor, J. J. (2019). An Engineered B. subtilis Inducible Promoter System with over 10 000-Fold Dynamic Range. ACS synthetic biology, 8(7), 1673–1678. https://doi.org/10.1021/acssynbio.8b00469

Design considerations: This part was codon optimized for expression in Bacillus subtilis. It produces the coenzyme for human pancreatic lipase which allows it to function in the presence of bile salts. Part BBa_ was designed to work in conjunction with this one.