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Lanmodulin with a C-terminus 6x His-Tag (LanM-His)

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.

Design

The lanmodulin sequence has been codon-optimized for expression in E. coli and with the addition of a 6x histidine tag to the C-terminus of the protein, it could easily be purified using the universal His-Tag purification protocol.

Sequence and Features

Characterization

We have successfully characterized lanmodulin’s REE binding capabilities. To do this we conducted a metal recovery assay where an initial sample of neodymium was incubated with lanmodulin. In this assay an initial known sample of neodymium was incubated with the protein. The protein fraction was then removed using centrifugal filtration and the filtrate was analyzed using the colorimetric Arsenazo assay.

Figure 1. Arsenazo III assay of samples before and after centrifugation. A starting neodymium sample of 25uM was treated with either water of 5uM lanmodulin. After centrifugal filtration, concentration of filtrate were assessed using Arsenazo III assay.

A darker color represents a high neodymium concentration while a lighter color represents a low neodymium concentration. After the addition of lanmodulin, the solution turned from a dark blue color to a light pink color. Thus, suggesting that lanmodulin was able to grab onto most of the neodymium.

Results from the Arsenazo was quantified by measuring absorbance at 650 nm. It was shown that the concentration of neodymium in the sample dropped from an initial value of 146 uM to 28 uM after incubation with lanmodulin, indicating an 81% neodymium recovery rate. Whereas the GST (negative control) left more than half of the neodymium in the solution, indicating that the observed neodymium binding capacity is unique to lanmodulin.

Figure 2. Concentration of neodymium in the initial sample, the filtrate and after treatment with HCl. The samples were treated with either 5 uM of lanmodulin, 5 uM of GST or water.

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Figure 3.Percent neodymium recovery efficiency of 5 uM lanmodulin, 5 uM GST and water from a 146 uM initial sample of neodymium.

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