Regulating Xylose Utilization By Disturbing Glucose Sensing System Of Saccharomyces Cerevisiae

The xylose metabolism capacity of recombinant Saccharomyces cerevisiae has been greatly improved through genetic engineering and adaptive evolution.However,the xylose consumption rate only approach to 50%of glucose consumption rate.Further improving the xylose utilization is important to realize the cost-effective production of the second-generation fuel ethanol.Our previous work showed that the extracellular glucose gave positive effect to xylose metabolism when the glucose could not enter the cell and did not inhibit the xylose absorption.This result indicated that the glucose sensoring system might regulated the xylose metabolism.It was reported that the xylose metabolism characteristics of S.cerevisiae is in the way between utilizing fermentation carbon source(glucose)and non-fermentation carbon source(glycerol,ethanol,acetate).Therefore,it is speculated that S.cerevisiae may sense xylose as semi-fermentative carbon source.There are two signal parthways in S.cerevisiae that sense the extracellular glucose signal.First,the Gprl,which does not sense xylose,sense the extracellular glucose signal to regulate the metabolism through the cAMP-PKA parthway.Second,the Snf3/Rgt2 senses the extracellular glucose signal to regulate the transcription of HXTs,HXK2,and et al.The respond mode of Snf3/Rgt2 to xylose is not clear.In presnt work,we investigated the effect of this two pathways to xylose metabolism by disturbing the proteins involved in.Among recombinant strains with GPA2G132V?RAS2G19V?pde1??pde1?pde2??pde?GPA2G132V,the genetic manipulation of GPA2G132V?pde1? pde2??pde1?AGPA2G132V increased the intracellular PKA activity and improved xylose utilization but decreased the growth.Expressing the GPA2G132V also improve the glucose utilization.On the contrary,RAS2G19V and pde1? have little effect on PKA activity,but also enhance xylose utilization.The alternation of transcriptional level of HXTs in strain with snf3?,rgt2?,and snf3?Argt2? revealed that both Snf3 and Rgt2 could respond to xylose,and the reponse related to the xylose level.When the concentration of xylose was low,the Snf3 and Rgt2 could not give signal to the downstream pathway.Moreover,we proved that konckouting the transcriptional repressor gene RGT1 increased the expression of HXTs therefore improve the xylose utilization.In conclusion,disturbing glucose sensing system could enhance sugar utilization of S.cerevisiae.The development of these regulation strategies and elements will bring positive effect on enhancing the xylose utilization of S.cerevisiae.