Part:BBa_K844002
Spider Silk 1x Subunit "W" (native sequence)
Spider silk 2E subunit native sequence; contains two elasticity domains and one strength domain.
IMPORTANT NOTE: This part uses Assembly Standard #23 (Silver Fusion) scar sites, which the sequence data below and on composite parts using this part will not reflect this scar sequence (it shows Assembly Standard #10 scars).
Usage and Biology
The sequence for this subunit was derived from the 2E silk subunit from Brooks et al. (2008), which is based on the native sequence of the MaSp2 silk gene in Argiope aurantia. The 2E silk subunit contains codons for only six amino acids in the following proportions: glycine (44.1%), glutamine (17.6%), proline (14.7%), alanine (11.8%), serine (7.4%), and tyrosine (4.4%). The native Argiope aurantia codon usage profile is significantly different from E. coli ’s and the GC% of the DNA molecule itself (75%) is significantly different from the average GC% of native E. coli genes (51%). These features can make the native form of the gene difficult and slow to produce for the ribosomes in the cell as the necessary tRNA molecules are present in lower concentrations in E. coli and the genetic machinery of E. coli is not adapted to opening DNA of high GC% when making mRNA, and thus lower amounts of spider silk mRNA are produced.
This part is designed for use as an internal subunit in a longer spider silk construct. This part should be preceded by the BBa_K844006 silk subunit, which is identical to this subunit but contains an added Met (ATG) start codon upstream, and should be followed by either more repeats of this subunit or end with a BBa_K844000 part, which contains a 10-Histidine tag and a double stop codon (TAATAA).
The silk subunit contains four main domains: two elasticity domains, a linker domain, and a strength domain. The elasticity domains contains sequences coding for beta-helices and beta-spiral in the protein; these structures increase the elasticity of the spider silk. The linker domain joins the strength and elasticity domains together. The strength domain will form beta-sheets and strengthen the silk fibers by cross-linking silk strands.
Silk Subunit Amino Acid Composition:
Related parts:
BBa_K844001 – the native spider silk subunit with one elasticity domains and one strength domain (no sequence manipulation)
BBa_K844003 – a codon optimized version of this part that uses only one codon for each amino acid
BBa_K844004 – a codon optimized version of this part that uses either one or two codons for each amino acid
BBa_K844006 – this spider silk subunit with an added Met (ATG) start codon at the 5’ end
BBa_K844016 - A codon optimized and improved concatemer based on K844002's sequence. BBa_K844016 has been shown to successfully produce spider silk protein and thread.
References:
Brooks AE, Stricker SM, Joshi SB, Kamerzell TJ, Middaugh CR, and Lewis RV. 2008. Properties of synthetic spider silk fibers based on Argiope aurantia MaSp2. Biomacromolecules 9:1506–10.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal NgoMIV site found at 19
Illegal NgoMIV site found at 103 - 1000COMPATIBLE WITH RFC[1000]
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