Team:Utah State/Project

Antimicrobial spider silk proteins (functionalized spider silk) could be used to make antimicrobial spider silk fibers or films. A previous study by Gomes et al. 2011 demonstrated that it is feasible to produce antimicrobial spider silk films. The Utah State iGEM team in 2012 was able to successfully produce spider silk proteins in E. coli with the use of BioBricks and successfully spin spider silk protein that was approximately 25.4 kDa in size. The parts that were submitted to the registry by Utah State in 2012 are RFC 23 compatible which allows for protein fusions, hence the addition of RFC 23 compatible AMPs to spider silk protein at the C or N terminal would be highly desirable.

The spider silk generator (BBa_K844016) created by Utah State in 2012 was able to produce proteins of 25.4 kDa. An increase of spider silk subunit repeats at the genetic level would enable a larger (and therefore ‘stronger’ fiber) to be produced. This year we proposed to double the number of spider silk repeat units to 8 and attach AMPs to either the C or N terminal ends of the spider silk. The purification of this antimicrobial functionalized silk would be performed with a 10x His-Tag fused to the spider silk, at the opposite terminus as the AMP. The possibility of "masking" the properties of the AMP were taken into account with our genetic design, which is why we did not create this fusion protein with the AMP located in between the spider silk and 10x His-Tag proteins. The figures below demonstrate the design of antimicrobial functionalized spider silk.

All constructs were assembled in pSB1C3 an E. coli as the chassis. AMPs were designed with RFC 23 under consideration to allow for protein fusions.