Part:BBa_K3945008

Lanmodulin with CipA cellulose binding domain (LanM-CipA)

Usage and Biology

Lanmodulin is recently a novel lanthanide binding protein from Methylobacterium extorquens that displays more than 100-million fold selectivity for rare earth elements [1]. Such affinity and selectivity have not been observed in any previously studied macromolecule. In addition, lanmodulin is extremely robust, capable of withstanding temperatures as high as 95 °C and pH levels as low as 2.5 [2]. Thus, allowing it to be the perfect molecular tool for us in an efficient rare earth recovery system.

A novel fusion protein with lanmodulin fused to the N-terminus of Clostridium thermocellum’s CipA cellulose binding module (CBMcipa). CBMcipa is a versatile and is able to from stable connections with both crystalline and regenerated amorphous cellulose. In addition, CBMcipa is one of the most characterized CBMs out there and has been shown to have a high efficiency of heterologous expression in E. coli, making an ideal choice for one step immobilization and purification of LanM from E. coli onto cellulose solid support [3].

Design

With a flexible and inert Threonine-Proline linker joining the two proteins, the immobilized lanmodulin will be able to operate freely on the surface of the cellulose support. The sequence of both proteins was also codon-optimized for expression in E. coli. A Factor Xa recognition site has been incorporated between the CBM and lanmodulin to cleave the lanmodulin protein free after purification on cellulose

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BglII site found at 701
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal NgoMIV site found at 88
  
• 1000
  COMPATIBLE WITH RFC[1000]

References

1. JA C, ER F, JA M, JV H, TN L. Lanmodulin: A Highly Selective Lanthanide-Binding Protein from a Lanthanide-Utilizing Bacterium. Journal of the American Chemical Society. 2018 [accessed 2021 Sep 17];140(44):15056–15061. https://pubmed.ncbi.nlm.nih.gov/30351021/. doi:10.1021/JACS.8B09842

2. GJ D, JA M, DM P, DW R, JA C, Y J. Selective and Efficient Biomacromolecular Extraction of Rare-Earth Elements using Lanmodulin. Inorganic chemistry. 2020 [accessed 2021 Sep 17];59(17):11855–11867. https://pubmed.ncbi.nlm.nih.gov/32686425/. doi:10.1021/ACS.INORGCHEM.0C01303

3.Improved immobilization of fusion proteins via cellulose‐binding domains - Linder - 1998 - Biotechnology and Bioengineering - Wiley Online Library. [accessed 2021 Oct 14]. https://onlinelibrary.wiley.com/doi/pdf/10.1002/(SICI)1097-0290(19981205)60:5%3C642::AID-BIT15%3E3.0.CO;2-8