Enzymes screening strategies

Fig. 1. Screening strategies to obtain new or improved enzymes by screening...

Based on the screening strategies discussed in Subheading 3.1., targeted colonies can be screened by PCR or Southern blot. Take 15 pL DNA for each restriction enzyme digestion, or, pool 5 samples of DNA to run PCR. The amount of DNA in each PCR reaction should be optimized with other reagents. 

Most recently, this group applied the metagenome screening strategy for fhe identification of improved DERA enzymes [75]. More than 15 DERA enzymes were discovered, of which one was used to optimize the process mentioned (Scheme 4.17E) on a 100 g scale. Using a fed-batch strategy, which is necessary because of substrate inhibition, Burk et al. obtained a final product concentration (16) of 93 g I. (>99.5% ee, de 96.6%). Concomitantly, the amount of enzyme was reduced to 2% (wt/wt). 

Coady TM, Coffey LV, O Reihy C, Owens EB, Lennon CM. A high throughput screening strategy for the assessment of nitrUe-hydrolyzing activity towards the production of enantiopure P-hydroxy acids.J Mol Catal B Enzym 2013 97 150-5. 

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