PEDS Advance Access originally published online on October 14, 2004
Protein Engineering Design and Selection 2004 17(9):673-680; doi:10.1093/protein/gzh079
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Stabilization of TRAIL, an all-ß-sheet multimeric protein, using computational redesign
1Department of Pharmaceutical Biology, University of Groningen, Antonius Deusinglaan 1, 9713 AV, Groningen, The Netherlands and 2Structural Biology and Biocomputing Program, EMBL, Meyerhofstrasse 1, D-69117 Heidelberg, Germany
3 To whom correspondence should be addressed. E-mail: w.j.quax{at}farm.rug.nl
Protein thermal stability is important for therapeutic proteins, both influencing the pharmacokinetic and pharmacodynamic properties and for stability during production and shelf-life of the final product. In this paper we show the redesign of a therapeutically interesting trimeric all-ß-sheet protein, the cytokine TRAIL (tumor necrosis factor-related apoptosis-inducing ligand), yielding variants with improved thermal stability. A combination of tumor necrosis factor (TNF) ligand family alignment information and the computational design algorithm PERLA was used to propose several mutants with improved thermal stability. The design was focused on non-conserved residues only, thus reducing the use of computational resources. Several of the proposed mutants showed a significant increase in thermal stability as experimentally monitored by far-UV circular dichroism thermal denaturation. Stabilization of the biologically active trimer was achieved by monomer subunit or monomer–monomer interface modifications. A double mutant showed an increase in apparent Tm of 8°C in comparison with wild-type TRAIL and remained biologically active after incubation at 73°C for 1 h. To our knowledge, this is the first study that improves the stability of a large multimeric ß-sheet protein structure by computational redesign. A similar approach can be used to alter the characteristics of other multimeric proteins, including other TNF ligand family members.
Received June 29, 2004; revised September 24, 2004; accepted October 5, 2004.
Edited by Jane Clarke
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