Role of xylose transporters in xylitol production from engineered Escherichia coli.
Escherichia coli W3110 was previously engineered to co-utilize glucose and xylose by replacing the wild-type crp gene with a crp* mutant encoding a cAMP-independent CRP variant (Cirino et al., 2006 [Cirino, P.C., Chin, J.W., Ingram, L.O., 2006. Engineering Escherichia coli for xylitol production from glucose-xylose mixtures. Biotechnol. Bioeng. 95, 1167-1176.]). Subsequent deletion of the xylB gene (encoding xylulokinase) and expression of xylose reductase from Candida boidinii (CbXR) resulted in a strain which produces xylitol from glucose-xylose mixtures. In this study we examine the contributions of the native E. coli xylose transporters (the d-xylose/proton symporter XylE and the d-xylose ABC transporter XylFGH) and CRP* to xylitol production in the presence of glucose and xylose. The final batch xylitol titer with strain PC09 (Delta xylB and crp*) is reduced by 40% upon deletion of xylG and by 60% upon deletion of both xyl transporters. Xylitol production by the wild-type strain (W3110) expressing CbXR is not reduced when xylE and xylG are deleted, demonstrating tight regulation of the xylose transporters by CRP and revealing significant secondary xylose transport. Finally, plasmid expression of XylE or XylFGH with CbXR in PC07 (Delta xylB and wild-type crp) growing on glucose results in xylitol titers similar to that achieved with PC09 and provides an alternative strategy to the use of CRP*.
Khankal
Reza
R
Department of Chemical Engineering, Pennsylvania State University, University Park, PA 16802, USA.
Chin
Jonathan W
JW
Cirino
Patrick C
PC
D018528:ATP-Binding Cassette Transporters / Q000235:genetics / Q000378:metabolism*; D003470:Culture Media; D002373:Cyclic AMP Receptor Protein / Q000235:genetics / Q000378:metabolism; D050542:D-Xylulose Reductase / Q000235:genetics / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism*; D017353:Gene Deletion; D024861:Genetic Enhancement; D005947:Glucose / Q000378:metabolism; D026901:Membrane Transport Proteins; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000235:genetics / Q000378:metabolism; D015202:Protein Engineering; D027981:Symporters / Q000235:genetics / Q000378:metabolism*; D014993:Xylitol / Q000096:biosynthesis*; D014994:Xylose / Q000378:metabolism
D018528:ATP-Binding Cassette Transporters; D003470:Culture Media; D002373:Cyclic AMP Receptor Protein; D029968:Escherichia coli Proteins; D026901:Membrane Transport Proteins; D027981:Symporters; C481662:crp protein, E coli; C033903:xylE protein, E coli; D014994:Xylose; D050542:D-Xylulose Reductase; D017853:Phosphotransferases (Alcohol Group Acceptor); C032700:xylulokinase; D005947:Glucose; D014993:Xylitol