Functional analysis of the Lactococcus lactis usp45 secretion signal in the secretion of a homologous proteinase and a heterologous alpha-amylase.
The ups45 gene encodes the major extracellular protein from Lactococcus lactis. The deduced sequence of the 27 residue leader peptide revealed the tripartite characteristics of a signal peptide. This leader peptide directed the efficient secretion of the homologous proteinase (PrtP) in L. lactis, indicating that the putative signal peptide of PrtP can be replaced by the 27 residue Usp45 leader peptide. In addition, the 27 residue leader peptide could be used to secrete the Bacillus stearothermophilus alpha-amylase, encoded by the amyS gene. Fusion of the usp45 promoter region and various parts of the leader sequence to an amyS gene devoid of its signal sequence, showed that in Escherichia coli the first 19, 20, and 27 residues of the Usp45 leader are able to direct alpha-amylase secretion. In L. lactis the shorter signal peptides did not result in secretion of alpha-amylase, providing experimental evidence for the hypothesis that gram-positive bacteria require a longer signal peptide for secretion than gram-negative organisms.
van Asseldonk
M
M
Department of Biophysical Chemistry, Netherlands Institute of Dairy Research (NIZO), Ede.
de Vos
W M
WM
Simons
G
G
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D001483:Base Sequence; D003001:Cloning, Molecular; D004274:DNA, Recombinant; D010450:Endopeptidases / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli; D001411:Geobacillus stearothermophilus / Q000201:enzymology; D013294:Lactococcus lactis / Q000201:enzymology / Q000378:metabolism*; D008969:Molecular Sequence Data; D011499:Protein Processing, Post-Translational; D021382:Protein Sorting Signals / Q000235:genetics* / Q000378:metabolism; D000516:alpha-Amylases / Q000235:genetics / Q000378:metabolism*
D001426:Bacterial Proteins; D004274:DNA, Recombinant; D021382:Protein Sorting Signals; C066583:usp45 protein, Lactococcus lactis; D000516:alpha-Amylases; D010450:Endopeptidases