tailieunhanh - Báo cáo khoa học: Increased NADPH concentration obtained by metabolic engineering of the pentose phosphate pathway in Aspergillus niger

Many biosynthetic reactions and bioconversions are limited by low availab-ility of NADPH. With the purpose of increasing the NADPH concentra-tion and⁄or the flux through the pentose phosphate pathway inAspergillus niger, the genes encoding glucose 6-phosphate dehydrogenase (gsdA), 6-phosphogluconate dehydrogenase (gndA) and transketolase (tktA) were cloned and overexpressed in separate strains. | ềFEBS Journal Increased NADPH concentration obtained by metabolic engineering of the pentose phosphate pathway in Aspergillus niger Bjarne R. Poulsen1 Jane N0hr2 Stephen Douthwaite2 Line V. Hansen2 Jens J. L. Iversen2 Jaap Visser1 and George J. G. Ruijter1 t 1 Molecular Genetics of IndustrialMicroorganisms Wageningen University the Netherlands 2 Department of Biochemistry and Molecular Biology University of Southern Denmark Odense Denmark Keywords Aspergillus niger gndA NADPH overexpression pentose phosphate pathway Correspondence . Iversen Department of Biochemistry and Molecular Biology University of Southern Denmark Campusvej55 DK-5230 Odense M Denmark Fax 45 6550 2467 Tel 45 6550 1000 ext 2376 E-mail jjli@ Present addresses FungalGenetics and Technology Consultancy 6700 AJ Wageningen the Netherlands tMetabolic Diseases Laboratory Leiden University MedicalCenter 2300 RC Leiden the Netherlands Database The nucleotide sequences reported are in the GenBank database under the accession numbers AJ551178 AJ551177 and AJ550995 Received 24 July 2004 revised 19 December 2004 accepted 4 January 2005 doi Many biosynthetic reactions and bioconversions are limited by low availability of NADPH. With the purpose of increasing the NADPH concentration and or the flux through the pentose phosphate pathway in Aspergillus niger the genes encoding glucose 6-phosphate dehydrogenase gsdA 6-phosphogluconate dehydrogenase gndA and transketolase tktA were cloned and overexpressed in separate strains. Intracellular NADPH concentration was increased two- to ninefold as a result of 13-fold overproduction of 6-phosphogluconate dehydrogenase. Although overproduction of glucose 6-phosphate dehydrogenase and transketolase changed the concentration of several metabolites it did not result in increased NADPH concentration. To establish the effects of overexpression of the three genes wild-type and overexpressing strains were characterized in detail