Purification and characterization of d-xylulokinase from the pentose-fermenting yeast Pichia stipitis NCYC 1541

Flanagan, Thomas R.; Waites, Matthew

doi:10.1016/0141-0229(92)90080-8

Cited by 29 publications

(30 citation statements)

References 17 publications

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“…The XYL3 sequence (GenBank accession no. AF127802 and AAF72328) from pX3 contained an open reading frame of 1,872 nucleotides, coding for a polypeptide of 623 amino acids with a total calculated molecular mass of 69,198 Da, which agreed well with the previous result (71 kDa) from a P. stipitis xylulokinase purification study (13). Sequence analysis of the 900 bp 5Ј to the open reading frame identified three putative TATA elements located at Ϫ180 to Ϫ188, Ϫ286 to Ϫ294, and Ϫ335 to Ϫ342.…”

Section: Strainssupporting

confidence: 81%

“…The D-xylulokinase from S. cerevisiae is also known to possess D-ribulokinase activity (31). The P. stipitis D-xylulokinase differs in that it does not possess Dribulokinase activity and hence is more specific for D-xylulose (13).…”

Section: Discussionmentioning

confidence: 99%

“…Thus, some controversy surrounds this approach. The S. cerevisiae D-xylulokinase is also active on D-ribulose (31), whereas the P. stipitis D-xylulokinase is not (13).…”

mentioning

confidence: 99%

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Molecular Cloning of XYL3 ( d -Xylulokinase) from Pichia stipitis and Characterization of Its Physiological Function

Liu¹,

Jones

Shi

et al. 2002

Appl Environ Microbiol

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XYL3, which encodes a D-xylulokinase (EC 2.7.1.17), was isolated from Pichia stipitis CBS 6054 genomic DNA by using primers designed against conserved motifs. Disruption of XYL3 eliminated D-xylulokinase activity, but D-ribulokinase activity was still present. Southern analysis of P. stipitis genomic DNA with XYL3 as a probe confirmed the disruption and did not reveal additional related genes. Disruption of XYL3 stopped ethanol production from xylose, but the resulting mutant still assimilated xylose slowly and formed xylitol and arabinitol. These results indicate that XYL3 is critical for ethanol production from xylose but that P. stipitis has another pathway for xylose assimilation. Expression of XYL3 using its P. stipitis promoter increased Saccharomyces cerevisiae D-xylulose consumption threefold and enabled the transformants to produce ethanol from a mixture of xylose and xylulose, whereas the parental strain only accumulated xylitol. In vitro, D-xylulokinase activity in recombinant S. cerevisiae was sixfold higher with a multicopy than with a single-copy XYL3 plasmid, but ethanol production decreased with increased copy number. These results confirmed the function of XYL3 in S. cerevisiae.

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Section: Strainssupporting

confidence: 81%

Section: Discussionmentioning

confidence: 99%

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Molecular Cloning of XYL3 ( d -Xylulokinase) from Pichia stipitis and Characterization of Its Physiological Function

Liu¹,

Jones

Shi

et al. 2002

Appl Environ Microbiol

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“…One di¡erence between the enzymes of the two organisms is that the P. stipitis enzyme is more speci¢c for D-xylulose, i.e. it has a much lower activity with D-ribulose than the S. cerevisiae enzyme [19].…”

Section: Xylulokinase Kineticsmentioning

confidence: 99%

The role of xylulokinase in Saccharomyces cerevisiae xylulose catabolism

Richard¹

2000

FEMS Microbiology Letters

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Many yeast species have growth rates on D-xylulose of 25^130% of those on glucose, but for Saccharomyces cerevisiae this ratio is only about 6%. The xylulokinase reaction has been proposed to be the rate-limiting step in the D-xylulose fermentation with S. cerevisiae. Overexpression of xylulokinase encoding XKS1 stimulated growth on D-xylulose in a S. cerevisiae strain to about 20% of the growth rate on glucose and deletion of the gene prevented growth on D-xylulose and D-xylulose metabolism. We have partially purified the xylulokinase and characterised its kinetic properties. It is reversible and will also accept D-ribulose as a substrate. ß

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“…Free D-xylose is found in guava, pears, blackberries, loganberries, raspberries, aloe vera gel, kelp, echinacea, boswellia, broccoli, spinach, eggplant, peas, green beans, okra, cabbage and corn. It is the second-most common saccharide in the natural world [1,2,4,13]. D-xylose undergoes differ- [6,7,8].…”

Section: Introductionmentioning

confidence: 99%

Cloning and Characterization of D-xylulose Kinase from Kocuria gwangalliensis Strain SJ2

Jeong¹,

Hwang²,

Seo³

et al. 2015

Journal of Life Science

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D-Xylulose is phosphorylated to D-xylulose-5-phosphate by D-xylulose kinase before it enters glycolysis via the nonoxidative pentose phosphate pathway. A gene encoding a novel D-xylulose kinase (XK) from K. gwangalliensis strain SJ2 was sequenced and expressed in E. coli. The sequence of the isolated XK gene was 1,419 bp, encoding 472 amino acids. The XK protein was more closely related to the Arthrobacter phenanthrenivorans XK than to the Bifidobacterium catenulatum one, as reflected in the sequence identity (54.9% vs. 38.7%). The XK gene was subcloned into the pCold-II expression vector. The resulting plasmid was transformed into E. coli strain BL21 (DE3) cells and the expression of the recombinant XK protein was induced by the addition of IPTG. The resulting protein was expressed as a fusion protein of approximately 48 kDa containing a N-terminal six-histidine extension that was derived from the expression vector. The expressed protein was homogenized by affinity chromatography and showed enzymatic activity corresponding to D-xylulose kinase. XK enzyme kinetic studies with D-xylulose and ATP showed a Km of 250±20 μM and 1,300±50 μM, respectively. The results obtained from this study will provide a wider knowledge base for the characterization of D-xylulose kinase at the molecular level.

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Purification and characterization of d-xylulokinase from the pentose-fermenting yeast Pichia stipitis NCYC 1541

Cited by 29 publications

References 17 publications

Molecular Cloning of XYL3 ( d -Xylulokinase) from Pichia stipitis and Characterization of Its Physiological Function

Molecular Cloning of XYL3 ( d -Xylulokinase) from Pichia stipitis and Characterization of Its Physiological Function

The role of xylulokinase in Saccharomyces cerevisiae xylulose catabolism

Cloning and Characterization of D-xylulose Kinase from Kocuria gwangalliensis Strain SJ2

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