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J. Biol. Chem., Vol. 279, Issue 46, 47890-47897, November 12, 2004

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TreT, a Novel Trehalose Glycosyltransferring Synthase of the Hyperthermophilic Archaeon Thermococcus litoralis^*

Qiuhao Qu, Sung-Jae Lee, and Winfried Boos

From the Department of Biology, University of Konstanz, 78457 Konstanz, Germany

The gene cluster in Thermococcus litoralis encoding a multicomponent and binding protein-dependent ABC transporter for trehalose and maltose contains an open reading frame of unknown function. We cloned this gene (now called treT), expressed it in Escherichia coli, purified the encoded protein, and identified it as an enzyme forming trehalose and ADP from ADP-glucose and glucose. The enzyme can also use UDP- and GDP-glucose but with less efficiency. The reaction is reversible, and ADP-glucose plus glucose can also be formed from trehalose and ADP. The rate of reaction and the equilibrium favor the formation of trehalose. At 90 °C, the optimal temperature for the enzymatic reaction, the half-maximal concentration of ADP-glucose at saturating glucose concentrations is 1.14 mM and the V_max is 160 units/mg protein. In the reverse reaction, the half-maximal concentration of trehalose at saturating ADP concentrations is 11.5 mM and the V_max was estimated to be 17 units/mg protein. Under non-denaturating in vitro conditions the enzyme behaves as a dimer of identical subunits of 48 kDa. As the transporter encoded in the same gene cluster, TreT is induced by trehalose and maltose in the growth medium.

Received for publication, May 4, 2004 , and in revised form, August 23, 2004.

^* This work was supported by grants from the Deutsche Forschungs-gemeinschaft and the Fonds der Chemischen Industrie. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

To whom correspondence should be addressed. Tel.: 0049-7531-882658; Fax: 0049-7531-883356; E-mail: winfried.boos{at}uni-konstanz.de.

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