Coding

Part:BBa_K3945005

Designed by: Ramin Kahidi   Group: iGEM21_Calgary   (2021-10-20)

Binding Affinity Optimized Mutated Lanmodulin I (BAO LanM-I)


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.

We have mutated the lanmodulin sequence using the hotspots mutagenesis model to improve the binding capacity of lanmodulin's fourth binding pocket.

Design

Likely modifications that improve protein stability were identified using HotSpot Wizard. These mutations were conducted to generate a large mutant library. Structural models were obtained to work with further. The members of this dictionary then were protonated at several different pH levels. The binding potential was then calculated for each pH level. The proteins with improved binding throughout the pH range were taken from the crop, and the rest were discarded. The two sequences with the highest binding potential throughout the range were selected to be shared so other researchers could utilize their improved binding and stability for different processes.

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BamHI site found at 111
    Illegal BamHI site found at 186
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 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

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