In order to determinate whether this rather moderate enhancement was due to cancelation between enhancing and deleterious effects of the different mutations, we determined the effect of the single mutations on Kemp eliminase activity. However, no strong cancellation was found (Table ). Overall, these results highlight the low efficiency and limited enhancements that are typical of non-focused library screening. There is little doubt, of course, that a directed evolution experiment would eventually lead to substantial enhancements in activity, but this will likely require many rounds of library preparation and screening, and also the focus of this study is the extent to which computational approaches can be used to enhance enzyme activity in lieu of (otherwise more costly) directed evolution experiments.