Part:BBa_K3381006

Improved Mst-CopC-CBM2a

Improved Mst-CopC-CBM2a is a fusion protein consisting of an improved Mst-CopC copper binding domain, as well as CBM2a, a cellulose-binding domain. A linker sequence that was found to naturally occur in a CBM2a fusion was used as a linker domain between Mst-CopC and CBM2a (Courtade, 2018). The CBM2a and flexible linker sequence was submitted as part BBa_K3381007. Additionally, a sequence for a His-tag as well as a TEV recognition sequence was added to the N-terminus for ease of purification (by immobilized-metal affinity chromatography [IMAC]).

Fusion proteins containing cellulose-binding modules (CBMs) have readily been used in industrial purification processes, where the CBMs act as affinity tags. Strikingly enough, waste processing seems to be an application that has not been as fervently explored (Zhou, 2020). Furthermore, current waste processing methods fail to provide a way to recover metal ions once extracted, leaving the playing field open to technologies that make metal recovery possible. This part’s CopC sequence has been engineered to improve copper binding. Compared to the original Mst-CopC (BBa_K3381002), the improved Mst-CopC has four mutations: Y34F, F3H, S81D, and H85D. These were selected using a rational protein design approach and validated using Rosetta protein modelling. These mutations were shown to result in a total interaction energy of -1698.64 kJ/mol, versus the -540.64 kJ/mol of the original CopC, a 3.14 fold improvement (see our engineering and model pages for more information).

Mst-CopC-CBM2a is a fusion protein that allows for extraction and recovery of copper(II) from aqueous waste. It binds irreversibly to cellulose, and can also bind Cu(II) (Koropatkin, 2007). This makes it appropriate for affinity chromatography or related uses.

Sequence and Features

Assembly Compatibility:

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

  Illegal NheI site found at 95
  Illegal NheI site found at 125
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BamHI site found at 785
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  COMPATIBLE WITH RFC[1000]