Test if gluten has been metabolized.

From 2012.igem.org

Home Team Official Team Profile Project Parts Submitted to the Registry Modeling Notebook Safety Sponsors Attributions

This page is for Steven's research.

http://eprints.uniss.it/5145/1/Forteschi_M_Study_of_peptidases_involved.pdf 33-mer peptide: LQLQPFPQPQLPYPQPQLPYPQPQLPYPQPQPF

http://www.sciencedirect.com/science/article/pii/0009898190900824 Detection and estimation of the barley prolamin content of beer and malt to assess their suitability for patients with coeliac disease

-a study confirming that beer contains levels of gluten antigens harmful to Celiacs.

http://www.sciencemag.org/content/297/5590/2275.full Structural Basis for Gluten Intolerance in Celiac Sprue

-The antigenicity of gluten seems to be due to proline and glutamine rich peptides, the most prevalent one being the 33-mer listed above. The sequence seems to be fairly conserved across barley and wheat.Three distinct patient-specific T cell epitopes identified previously in T cell proliferation assays are present in this peptide, namely, PFPQPQLPY, PQPQLPYPQ (three copies), and PYPQPQLPY (two copies).

http://www.sunrisescience.com/Files/pTEF-MF.pdf pTEF-MF constitutive yeast vector marked for secretion

-MF-alpha secretion tag for yeast is used in this plasmid. I used the plasmid map to get the tag's sequence and BLASTed it. A 100% match confirmed that the sequence is present in yeast. We will attach this to the N-terminus of Kumamolisin, hopefully enabling the protein to be secreted in yeast.

http://www.jbc.org/content/263/13/6209.full.pdf Prepro-a-factor Has a Cleavable Signal Sequence

-MF-alpha signal sequence is attached to the animo-terminus of MF-alpha and cleaved in the golgi.

http://synbio.org.uk/dna-assembly/guidetogibsonassembly.html Gibson assembly

http://www.ncbi.nlm.nih.gov/pmc/articles/PMC391546/pdf/pnas00616-0033.pdf Alphafactor- directed synthesis and secretion of mature foreign proteins in Saccharomyces cerevisiae

-The peptide mating pheromone alpha-factor is efficiently secreted into the medium. Plasmids with the secretion factor and foreign proteins were transformed into yeast. Yeast cells transformed with some of these plasmids efficiently synthesize, process, and secrete into the medium the mature biologically active foreign proteins.

http://www.springerlink.com/content/l2115622t21t728w/fulltext.pdf Engineering of protein secretion in yeast: strategies and impact on protein production

http://ajpgi.physiology.org/content/291/4/G621.long Highly efficient gluten degradation with a newly identified prolyl endoprotease: implications for celiac disease

http://onlinelibrary.wiley.com/doi/10.1002/jms.1667/full Identification of N-glycosylation in prolyl endoprotease from Aspergillus niger and evaluation of the enzyme for its possible application in proteomics

http://mcl1.ncifcrf.gov/wlo_pubs/261.pdf enzyme structure and kinetics for Kumamolisin

http://www.jbc.org/content/282/36/26401.full.pdf+html Structure and function of EnvZ, specifically looking at the histidine kinase domain and the HAMP domain.