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Journal of Bacteriology, June 2001, p. 3428-3435, Vol. 183, No. 11
0021-9193/01/$04.00+0 DOI: 10.1128/JB.183.11.3428-3435.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.
Novel Type of Glucose-6-Phosphate Isomerase in the
Hyperthermophilic Archaeon Pyrococcus furiosus
Thomas
Hansen,
Margitta
Oehlmann, and
Peter
Schönheit*
Institut für Allgemeine Mikrobiologie,
Christian-Albrechts-Universität Kiel, D-24118 Kiel, Germany
Received 11 December 2000/Accepted 5 March 2001
Glucose-6-phosphate isomerase (phosphoglucose isomerase [PGI])
(EC 5.3.1.9) from the hyperthermophilic archaeon Pyrococcus furiosus was purified 500-fold to homogeneity. The enzyme had an
apparent molecular mass of 43 kDa and was composed of a single type of
subunit of 23 kDa indicating a homodimeric (
2)
structure. Kinetic constants of the enzyme were determined at the
optimal pH 7 and at 80°C. Rate dependence on both substrates followed Michaelis-Menten kinetics. The apparent Km
values for glucose-6-phosphate and fructose-6-phosphate were 8.7 and 1.0 mM, respectively, and the corresponding apparent
Vmax values were 800 and 130 U/mg. The enzyme
had a temperature optimum of 96°C and showed a significant thermostability up to 100°C, which is in accordance with its
physiological function under hyperthermophilic conditions. Based on the
N-terminal amino acid sequence of the subunit, a single open reading
frame (ORF; Pf_209264) was identified in the genome of P. furiosus. The ORF was characterized by functional overexpression
in Escherichia coli as a gene, pgi, encoding
glucose-6-phosphate isomerase. The recombinant PGI was purified and
showed molecular and kinetic properties almost identical to those of
the native PGI purified from P. furiosus. The deduced amino
acid sequence of P. furiosus PGI did not reveal significant
similarity to the conserved PGI superfamily of eubacteria and eucarya.
This is the first description of an archaeal PGI, which represents a
novel type of PGI.
*
Corresponding author. Mailing address: Institut
für Allgemeine Mikrobiologie,
Christian-Albrechts-Universität Kiel, Am Botanischen Garten 1-9, D-24118 Kiel, Germany. Phone: 49-431-880-4328. Fax: 49-431-880-2194. E-mail: peter.schoenheit{at}ifam.uni-kiel.de.
Journal of Bacteriology, June 2001, p. 3428-3435, Vol. 183, No. 11
0021-9193/01/$04.00+0 DOI: 10.1128/JB.183.11.3428-3435.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.
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