Mh. Metzger et Cp. Hollenberg, ISOLATION AND CHARACTERIZATION OF THE PICHIA-STIPITIS TRANSKETOLASE GENE AND EXPRESSION IN A XYLOSE UTILIZING SACCHAROMYCES-CEREVISIAE TRANSFORMANT, Applied microbiology and biotechnology, 42(2-3), 1994, pp. 319-325
The gene TKT from P. stipitis, encoding the enzyme transketolase (EC 2
.2.1.1), was cloned from a genomic library by hybridization with a S.
cerevisiae TKT1-gene-specific probe. The nucleotide sequence determine
d contains an open-reading frame of 2085 base pairs (bp) encoding a pr
otein of 695 amino acids with a predicted molecular mass of 75 113 Da.
The TKT gene was actively expressed in S. cerevisiae when placed unde
r the control of the homologous PDC1(- 15) promoter and could compleme
nt a S. cerevisiae tkt deletion. The TKT protein was immunologically d
etectable using S. cerevisiae transketolase-specific antiserum. Overex
pression of the P. stipitis TKT gene in a xylose-utilizing S. cerevisi
ae XYL1/XYL2 integrant led to a drastically extended generation time d
uring growth on xylose minimal medium under aerobic conditions.