초록
<P><B>Abstract</B></P><P><B>BACKGROUND:</B> <I>Yarrowia lipolytica</I> lipase LIP2 (YlLIP2) is an important industrial enzyme that has many potential applications. Although it has been successfully expressed in <I>Pichia pastoris</I> under the control of the <I>AOX1</I> promoter (<I>pAOX1</I>), there have been many efforts to develop new alternative promoters to <I>pAOX1</I> in order to avoid using methanol in the fermentation. Investigation of YlLIP2 production in <I>P. pastoris</I> using the formaldehyde dehydrogenase 1 promoter (<I>pFLD1</I>) is especially attractive, since little is known about its application in methanol‐free culture strategies.</P><P><B>RESULTS:</B> Three fed‐batch cultivations were performed to investigate the production of YlLIP2 in a <I>pFLD1</I>‐based system. When methanol was used as the fed‐batch feeding substrate, the maximum YlLIP2 activity obtained in a 10‐L bioreactor was 30 000 U mL<SUP>−1</SUP> after 143 h of culture, whereas the maximum YlLIP2 activity was further increased to 35 000 U mL<SUP>−1</SUP> by adopting a co‐induction strategy with methanol and methylamine as a mixed fed‐batch substrate. Furthermore, the maximum YlLIP2 activity reached 13 000 U mL<SUP>−1</SUP> after 80 h of cultivation in a methanol‐free culture.</P><P><B>CONCLUSION:</B> The expression levels of YlLIP2 in the <I>pFLD1</I>‐based system were comparable with those in a <I>pAOX1</I>‐based system. The results suggest that <I>pFLD1</I> is an attractive alternative to <I>pAOX1</I>, and may make it feasible to induce high yields of protein expression. Copyright © 2011 Society of Chemical Industry</P>