Fig. 2

Identification of LipA variants with enhanced extracellular activity or amount of protein. a Schematic representation of the two-step screening procedure. In step I, 29,199 LipA clones were analyzed for increased extracellular lipase activity in the culture supernatant of B. subtilis TEB1030. 175 clones were sequenced and 80 LipA variants identified with increased extracellular lipase activity (see panel b). In a second step, culture supernatants of these variants were analyzed as nine biological replicates for increased extracellular lipase activity and protein amount. b 80 LipA clones with increased extracellular lipase activity. The relative extracellular lipase activity of the LipA clones is plotted against the substituted amino acid position. c LipA variants with increased extracellular lipase amount. The extracellular lipase amount is plotted against the substituted amino acid position. In panels b and c, each black dot represents one LipA variant, and the grey bars mark the highly conserved amino acid positions (≥ 95%). Wild-type values, which were 0.57 ± 0.12 U/ml and 3.7 ± 0.6 µg/ml, respectively, were set to 1 (wtLipA) and the grey horizontal dotted lines mark the wtLipA standard deviation (σ)