초록
<P>Abstract</P><P>The hyperthermophilic α-amylase from <I>Thermococcus</I> sp. HJ21 possesses unique traits (Ca<SUP>2+</SUP>-independent thermostability and optimal temperature of 95°C) that make it a great potential candidate for use in the food industry. However, this Archaea isolated from a deep-sea thermal vent requires strict control of culture conditions and produces only small amounts of α-amylase. To solve these problems, the α-amylase gene was cloned and expressed in <I>Bacillus subtilis</I>, which is an ideal food-grade host for heterologous protein expression. To express high levels of this α-amylase in <I>B. subtilis</I>, the promoters P<SUB><I>grac</I></SUB>, P<SUB><I>xylA</I></SUB>, P43, and P<SUB><I>hag</I></SUB> were used to construct four different expression vectors for testing. The vector containing the P<SUB><I>xylA</I></SUB> promoter was found to have the highest transcriptional activity and produce the highest amylase activity (19.6 U/ml). To test the secretion efficiency of signal peptides in <I>B. subtilis</I>, three signal peptides were cloned and fused to the α-amylase gene (lacking its native signal peptide). The optimal signal peptide was S<SUB><I>amyQ</I></SUB>, with a secretion efficiency of approximately 90%. These results indicate that the promoter P<SUB><I>xylA</I></SUB> and signal peptide S<SUB><I>amyQ</I></SUB> tested in this study may be useful for the expression and secretion of archaeal proteins in <I>B. subtilis</I>.</P><P>Copyright © 2013 S. Karger AG, Basel</P>