Part:BBa_K1947029

BBa_K1947029

Usage and Biology

Protein purification is commonly used in Biochemical research, but can become a very tedious and inefficient procedure. We developed a system that can be used to conveniently and efficiently purify recombinant proteins with help of magnetosome. The system consists of two parts. First, we used E. coli to express an interest protein tagged by a Spytag. Second, we generated a Spycatcher-fused Mms13 protein and expressed it in the magetotactic bacteria AMB-1. These bacteria synthesize magnetosomes covered by phospholipid bilayer membrane, in which Mms13 is tightly anchored. Spycatcher binds Spytag with high affinity, and thus Spycatcher-Mms13 anchored in magnetosomes can strongly bind Spytag-conjugated protein and specifically bring it down with help of magnetic field. Our system is applicable to efficiently purifying any interest protein for different research purposes.

TEV protease may bring some impure proteins. We use intein to replace the TEV site. The M. xenopi GyrA intein provides a paradigm for a minimal protein splicing element. Intein can be cleaved by DTT. DTT as reducing agent protects the free thiol groups of the protein from oxidation and so it doesn’t bring foreign substance. As a result, intein can achieve a better effect of purification than TEV protease.

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal NgoMIV site found at 1230
  
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI.rc site found at 1150