Evolutionary protein stabilization in comparison with computational design

Two major strategies are currently used for stabilizing proteins: in vitro evolution and computational design. Here, we used gene libraries of the 1 domain of the streptococcal protein G (G beta 1) and Proside, an in vitro selection method, to identify stabilized variants of this protein. In the G beta 1 libraries, the codons for the four boundary positions 16, 18, 25, and 29 were randomized. Many G beta 1 variants with strongly increased thermal stabilities were found in 11 selections performed with five independent libraries. Previously, Mayo and co-workers used computational design to stabilize G beta 1 by sequence optimization at the same positions. Their best variant ranked third within the panel of the selected variants. None of the ten computed sequences was found in the Proside selections, because several computed residues for positions 18 and 29 were not optimal for stability. (c) 2005 Elsevier Ltd. All rights reserved.