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Title:   Author  item
                                            Hughes, Stephenitem
                                            Jackson Jr, Johnitem
                                            Dowd, Patrickitem
                                            Hector, Ronald

                                                         - *Ron*item
                                            Bischoff, Kennethitem
                                            STERNER, DAVID - PROGENRA, INC.item
                                            BANG, SOOKIE - SDSM&Titem
                                            GRYNAVYSKI, NICOLE - ARRYX INCitem
                                            CHAKRABARTY, TANIA - ARRYX INCitem
                                            Johnson, Ericitem
                                            LI, XIN-LIANG - TACOMA, WASHINGTONitem
                                            CAUGHEY, ROBERT - U OF I MED SCHOOL/PEOitem
                                            Liu, Siqingitem
                                            Skory, Christopher

                                                         - *Chris*item
                                            BUTT, TAUSEEF - LIFESENSORSitem
                                            LABAER, JOSHUA - HARVARD INSTITUTEitem
                                            Dien, Bruceitem
                                            Saha, Badalitem
                                            BOWEN, MICHAEL - ARS, FBTitem
                                            Qureshi, Nasibitem
                                            Rich, Josephitem
                                            Leathers, Timothyitem
                                            Cotta, Michaelitem
                                            FARRELLY, PHIL - HUDSON CONTROLitem
                                            Slininger, Patricia

                                                         - *Pat*
  Submitted to: Meeting Abstract
                                    Publication Type: Abstract Only
                                    Publication Acceptance Date: 5/16/2008
                                    Publication  Date: 5/16/2008
                                    Citation: Hughes, S.R., Jackson Jr, J.S., Dowd, P.F., Hector, R.E., Bischoff, K.M., Sterner, D., Bang, S., Grynavyski, N., Chakrabarty, T., Johnson, E.T., Li, X., Caughey, R.J., Liu, S., Skory, C.D., Butt, T., Labaer, J., Dien, B.S., Saha, B.C., Bowen, M., Qureshi, N., Rich, J.O., Leathers, T.D., Cotta, M.A., Farrelly, P., Slininger P.J. 2008. Three-vector system for high-level functional expression of value-added co-products with Xylose Isomerase and Xylulokinase in an industrial Saccharomyces cerevisiae strain [abstract]. Cambridge Healthtech Institute, Second Generation Biofuels Development Summit. Poster #1. p. 4.Interpretive Summary:
                                Technical Abstract: The three-plasmid yeast expression system utilizing the portable small ubiquitin-like modifier (SUMO) vector set combined with the efficient endogenous yeast protease rapidly produces large amounts of soluble functional protein.  It provides high levels of expression for three different proteins simultaneously.  This system is used to express a peptide of potential commercial value in an industrial Saccharomyces cerevisiae strain also engineered to express xylose utilization and metabolic correction enzymes.  The xylose isomerase (XI) gene for pentose utilization is expressed using the first of three SUMO vectors.  A value-added co-product is expressed in the second vector.  The test product here is the putative bioinsecticidal peptide, lycotoxin-1 (Lyt-1).  The third vector is employed to express metabolic correction genes to enhance pentose utilization.  Engineering of the yeast strain involved three steps.  First, using a novel PCR assembly strategy we cloned the XI open reading frame (ORF) into the URA-selectable SUMO vector and placed it into the S. cerevisiae strain.  Second, using amino acid scanning mutagenesis we generated a library of mutagenized Lyt-1 ORFs, cloned it into the TRP-selectable SUMO vector, and placed it into the XI-yeast.  Third, the xylulokinase gene was cloned into the LEU-selectable SUMO vector and placed into the Lyt-1-XI-yeast.  Yeast strains expressing xylulokinase and Lyt-1 in addition to XI showed improved growth on xylose compared to XI-yeast.


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