Expression of a wheat alpha-amylase gene in Escherichia coli: recognition of the translational initiation site and the signal peptide.

Transcription of a full-length cDNA clone of wheat alpha-amylase using a lac promoter in Escherichia coli results in synthesis of a precursor alpha-amylase polypeptide of the correct size, indicating that translation initiates correctly. Recognition of the plant translational initiation site by E. coli ribosomes is 15-20% as efficient as the ribosome-binding site of the beta-lactamase gene when it is fused to alpha-amylase. The alpha-amylase signal peptide is recognised in E. coli resulting in secretion of the enzyme into the periplasmic space; deletion of the signal peptide prevents secretion. Replacement of the alpha-amylase signal peptide with a beta-lactamase signal peptide also enables the bacterial cell to secrete the enzyme. The presence of the beta-lactamase and the alpha-amylase signal peptides in tandem results in secretion of the enzyme and removal of both signal peptides.