EXPRESSION AND SECRETION OF A XYLANASE FROM THE EXTREME THERMOPHILE, THERMOTOGA STRAIN FJSS3B.1, IN KLUYVEROMYCES-LACTIS

The yeast Kluyveromyces lactis has been developed as a host for extrac ellular production of thermophilic hemicellulases by employing express ion vectors based on the 2 mu-like plasmid pKD1 of Kluyveromyces droso philarium. A beta-1,4-xylanase gene (xynA) from the extreme thermophil e Thermotoga sp. strain FjSS3B.1 was fused in-frame with a synthetic s ecretion signal derived from the K. lactis killer toxin and expressed under control of the K. lactis LAC4 (beta-galactosidase) promoter, Cor rectly processed xylanase enzyme with full biological activity on oat spelts xylan was secreted during shake-flask cultivation of K. lactis transformants. The transcriptional activity of the LAC4 promoter drama tically affected mitotic stability of the expression vector under nons elective conditions. However, one combination of host strain and expre ssion plasmid showed higher stability and good yield and has been empl oyed for scaled-up production of XynA and other thermostable hemicellu lases in chemostat culture. XynA secreted by K. lactis is as thermosta ble as the native enzyme, having a half-life of 48 h at 90 degrees C.