Part:BBa_K1763000

Apis mellifera (honeybee) silk fibroin 3

This is the coding region for the honeybee silk protein #3. Previous literature has shown that it is possible to generate honeybee fibers by expressing this protein recombinantly in e. coli and spinning the protein into fibers in vitro It does not contain any of the regulatory elements necessary for protein expression.

Usage and Biology

Because this is just the coding region for the honeybee silk sequence, it is not meant to be expressed on its own. For an expressible honey bee silk biobrick part, please see BBa_K1763007 . Silk from Apis Mellifera represents an intriguing alternative to silks from spiders or silkworms. Although it is not quite as strong as these other types of silks, working with honey bee silk has certain advantages over spider and silkworm silk. The size of the honey bee silk protein gene is considerably smaller than the silk genes of spiders or silkworms. More importantly, the gene sequence is non repetitive, which allows us to synthesize and make modifications to the gene without the complications that are inherent to repetitive DNA sequences (citation). Honey bee silk also has a very different secondary and tertiary structure than spider and silkworm silks. It forms primary alpha helices, and four silk proteins come together to form a coiled coil structure these coiled coils are formed from four similar, yet unique proteins, Amelf 1-4. However, a study has shown that using one of these proteins, (Amelf3) is sufficient to reproduce the physical properties of the wild type fibers

Characterization

1. Protein Expression

• Here is the SDS gel of our expression of the honeybee silk protein. It was purified using [http://wolfson.huji.ac.il/purification/PDF/Protein_Expression_Extraction/NOVAGEN_BugBuster_protein_extraction.pdf this inclusion body protocol]

Fig. 1 Expected size of product is 40.0 kDA

• There are contaminating bands present on the SDS PAGE gel, indicating that the purification is not completely effective.
• We also ran BCA protein concentration assays on our purified honeybee proteins. Based on this assay we were able to get around 12 mg of protein from a 300 ml cell culture.
  • However, this 12 mg might not be composed entirely of honeybee protein because based on the SDS PAGE results, the purification was not 100% complete.

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
  COMPATIBLE WITH RFC[25]
• 1000
  COMPATIBLE WITH RFC[1000]