3.1.3.72 | synthesis |
transgenic expression in Pichia pastoris. Increasing isoform Alp2 copy number is detrimental to heterologous expression, clones with one copy of wild-type Alp2 produce the highest activity, clones with two, four and seven or more copies produce 70%, 25% and 10% respectively, of enzyme activity. Use of a sequence-optimized Alp2 gene increases the yield of the active enzyme by 25-50% in one/two copy clones. Reducing the temperature during heterologous expression leads to increases of 1.2-20fold suggesting that protein folding and post-translational processes may be the dominant factors limiting ALP2 expression |
730874 |