Team:WashU StLouis/Improve

Part BBa_K2663005: Aga2 and (Glycine3Ser)4 Linker

We decided to improve part BBa_K416004 designed by the 2010 NYU iGEM team. This composite part contained the Aga2 yeast surface display protein linked at the C-terminus to a flexible (Gly3Ser)4 linker. It can be used to attach to Aga1 on the surface of Saccharomyces cerevisiae EBY100. The original part was assembled by combining Aga2 with the (Gly3Ser)4 linker using RFC 10 which can create out-of-frame fusions. Since Aga2, the linker, and the wanted protein must all be encoded as one transcript and combined amino acid sequence, maintaining the open reading frame is crucial.

To fix these errors, our team redesigned the part by fixing the out of frame design and attached a yeGFP to the linker (designated as BBa_K2663005). Since Aga1 is controlled by the Gal1 promoter in EBY100, we found it to be convenient to also control the Aga2 construct with the Gal1 promoter. For the convenience of future use, we have submitted the full part with a Gal1 promoter attached (BBa_K2663006) to the registry. Any terminator can be used with the part.