Crystal structure of heat-labile enterotoxin from Escherichia coli with increased thermostability introduced by an engineered disulfide bond in the A subunit

Protein Science : a Publication of the Protein Society
F van den AkkerW G Hol

Abstract

Cholera toxin (CT) produced by Vibrio cholerae and heat-labile enterotoxin (LT-I), produced by enterotoxigenic Escherichia coli, are AB5 heterohexamers with an ADP-ribosylating A subunit and a GM1 receptor binding B pentamer. These toxins are among the most potent mucosal adjuvants known and, hence, are of interest both for the development of anti-diarrheal vaccines against cholera or enterotoxigenic Escherichia coli diarrhea and also for vaccines in general. However, the A subunits of CT and LT-I are known to be relatively temperature sensitive. To improve the thermostability of LT-I an additional disulfide bond was introduced in the A1 subunit by means of the double mutation N40C and G166C. The crystal structure of this double mutant of LT-I has been determined to 2.0 A resolution. The protein structure of the N40C/G166C double mutant is very similar to the native structure except for a few local shifts near the new disulfide bond. The introduction of this additional disulfide bond increases the thermal stability of the A subunit of LT-I by 6 degrees C. The enhancement in thermostability could make this disulfide bond variant of LT-I of considerable interest for the design of enterotoxin-based vaccines.

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Citations

May 4, 2000·Protein Science : a Publication of the Protein Society·R E BurtonT G Oas
Apr 5, 2000·The Journal of Biological Chemistry·T A Klink, R T Raines
Jun 25, 2004·Protein Science : a Publication of the Protein Society·Robert Schultz-HeienbrokNorbert Sträter
Jun 30, 2006·Proteins·Jean-Luc Pellequer, Shu-wen W Chen
Mar 17, 2021·Expert Review of Vaccines·Yizhi Qi, Christopher B Fox

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