XJTLU-CHINA-2023 has developed a total of eighteen distinct parts including nine new basic parts, nine new composite parts and five new improved parts. These components have been devised to tackle the issue of heavy and noble metal pollution within industrial wastewater. Below, we provide a concise overview and explanation of these three categories of components. For more in-depth details, please refer to the respective sections.
To assess the performance of the two fusion proteins we've generated, we conducted evaluations of their capacity to adsorb heavy metal ions and noble metal ions in wastewater.
For the New Basic Parts, CBM (BBa_K4968002), sfGFP (BBa_K4968003), and pUC_GW_Kan R (BBa_K4968012) as integral elements for protein expression. These elements are designed to adsorb heavy metal ions such as cadmium, copper, and lead. Additionally, codon-optimized CsgA (BBa_K4968004) and AG4 (BBa_K4968005) will serve as fundamental building blocks for protein expression with the capability to adsorb the noble metal ion silver. Furthermore, we've introduced codon-optimized SUMO (BBa_K4968007) tags to enhance the solubility of the target protein.
Within the New Composite Parts, CsgC-CsgD (BBa_K4968009), CsgE-CsgF-CsgG (BBa_K4968010), and CsgA-AG4-CsgC-CsgD-CsgE-CsgF-CsgG (BBa_K4968011) as composite components, designed to adsorb noble metal ions.
Similarly, we've combined CBM-sfGFP-pUC_GW_Kan R-SUMO (BBa_K4968017), SmtA-CBM-sfGFP (BBa_K4968018), SUMO-CBM-sfGFP (BBa_K4968019), MSmtA4-CBM-sfGFP (BBa_K4968020), and SUMO-MSmtA4-CBM-sfGFP (BBa_K4968021) to create composite protein units with heavy metal ion adsorption capabilities.
In the realm of New Improved Parts. Furthermore, we've developed SUMO-SmtA-CBM-sfGFP (BBa_K4968019), MSmtA4-CBM-sfGFP (BBa_K4968020), and SUMO-MSmtA4-CBM-sfGFP (BBa_K4968021) as enhancement components, all geared towards enhancing the adsorption of heavy metal ions. Additionally, RecomSwapNeo R/Kan R (BBa_K4968006) plays a pivotal role as a crucial fragment for knocking out CsgA-CsgB in MC4100 and BL21(DE3) genomes.
For comprehensive insights into the functionality and implementation of these components, please consult the dedicated sections elaborating on each one.
| Type | Part Number | Name | Length | Description |
|---|---|---|---|---|
| Basic | MSmtA4 | 168 bp | MSmtA4, a mutant protein derived from SmtA (BBa_K4968001) with four mutated sites and codon optimization, effectively enhances the adsorption capacity of heavy metals | |
| Basic | Optimized SmtA | 168 bp | SmtA, a protein belonging to the metallothionein family, and it has the ability to adsorb heavy metals and chelate these heavy metals in an inactive form. | |
| Basic | Optimized CBM | 315 bp | Carbohydrate Binding Modules (CBMs) are components of several enzymes, and their primary function is to bind to specific carbohydrates. | |
| Basic | Optimized sfGFP | 711 bp | sfGFP, a fluorescent labeling tool, does not lead to misfolding when fused with other proteins, thereby enhancing the stability of the fused protein. | |
| Baisc | Optimized CsgA | 453 bp | CsgA, a gene has remarkable ability to self-assemble into curli fiber amyloid nanofibers, optimized through codon optimization. | |
| Basic | Optimized AG4 | 39 bp | AG4, a synthetic silver-binding dodecapeptide, optimized through codon optimization. | |
| Basic | RecomSwapNeo R/Kan R | 1528 bp | The improved part of Low to medium copy Lambda Red recombineering compatible plasmid (BBa_K592202). As a Vector for Lambda Red Homologous Recombination. | |
| Tag | Optimized SUMO tag | 294 bp | Small ubiquitin-like modifiers (SUMOs) are widely used as tags for protein expression and purification. | |
| Composite | CsgA-AG4 | 510 bp | This composite part combines Optimized CsgA (BBa_K4968004) and AG4 (BBa_K4968005), containing a 6 amino acid linker. | |
| Composite | CsgC-CsgD | 1022 bp | This composite part combines CsgC (BBa_K1583001) and CsgD (BBa_K805015), containing a 38 bp nonsense sequence. | |
| Composite | CsgE-CsgF-CsgG | 1717 bp | This composite part combines CsgE (BBa_K4161013), CsgF (BBa_K4161014), and CsgG (BBa_K4161015), containing two 38 bp nonsense sequences. | |
| Composite | CsgA-Ag4-CsgC-CsgD-CsgE-CsgF-CsgG | 3293 bp | This composite part combines CsgA-AG4 (BBa_49680008), CsgC-CsgD (BBa_K4968009), and CsgE-CsgF-CsgG (BBa_K4968010), containing two 38 bp nonsense sequences. | |
| Basic | pUC_GW_Kan R | 2626 bp | As a Plasmid backbone for the part CBM-sfGFP-pUC_GW_Kan R-SUMO (BBa_K4968017). | |
| Composite | CBM-sfGFP-pUC_GW_Kan R-SUMO | 3991 bp | This composite part combines CBM (BBa_49680002), sfGFP (BBa_49680003), pUC_GW_Kan R (BBa_49680012), and SUMO (BBa_49680007). As a Vector for Gibson assembly with MSmtA4 (BBa_K4968000). | |
| Composite | SmtA-CBM-sfGFP | 1242 bp | This composite part combines SmtA (BBa_K4968001), CBM (BBa_K4968002), and sfGFP (BBa_K4968003), containing two 8 amino acid flexible linker. | |
| Composite | SUMO-SmtA-CBM-sfGFP | 1554 bp | This composite part combines SmtA (BBa_K4968001), CBM (BBa_K4968002), sfGFP (BBa_K4968003), and SUMO (BBa_K4968007), containing a 6 amino acid flexible linker and two 8 amino acid flexible linker. | |
| Composite | MSmtA4-CBM-sfGFP | 1242 bp | This composite part combines MSmtA4 (BBa_K4968000), CBM (BBa_K4968002), and sfGFP (BBa_K4968003), containing two 8 amino acid flexible linker. | |
| Composite | SUMO-MSmtA4-CBM-sfGFP | 1554 bp | This composite part combines MSmtA4 (BBa_K4968000), CBM (BBa_K4968002), sfGFP (BBa_K4968003), and SUMO (BBa_K4968007), containing a 6 amino acid flexible linker and two 8 amino acid flexible linker. |