Foreign signal peptides can constitute a barrier to functional expression of periplasmic proteins in Haemophilus influenzae.
To study the periplasmic branch of iron (ferric ion) uptake systems in Gram-negative bacteria, genetic reconstitution experiments were initiated in Haemophilus influenzae involving exchange of the periplasmic iron-binding protein. The expression of many of the heterologous periplasmic ferric-binding proteins (FbpAs) was quite limited. Transformation experiments with the fbpA gene from Neisseria gonorrhoeae yielded two colony sizes with different phenotypic characteristics. The small colonies contained the intact N. gonorrhoeae fbpA gene and were deficient in utilization of transferrin iron. The large colonies contained hybrid H. influenzae/N. gonorrhoeae fbpA genes, were proficient in transferrin iron utilization and had enhanced levels of expression of FbpA. These hybrid genes included several that encoded the mature N. gonorrhoeae FbpA with the H. influenzae signal peptide. To more fully evaluate the effect of foreign signal peptides, a series of hybrid genes were prepared that exchanged the signal peptides from H. influenzae FbpA, N. gonorrhoeae FbpA and the TEM-1 beta-lactamase. The presence of the H. influenzae leader was required for functional expression of FbpAs and was shown to dramatically increase the level of beta-lactamase activity.
Department of Microbiology and Infectious Diseases, Faculty of Medicine, University of Calgary, Heritage Medical Research Building, 3300 Hospital Dr. NW, Calgary, Alberta, Canada T2N 4N1.
Siu Ling
SL
Lam
Department of Microbiology and Infectious Diseases, Faculty of Medicine, University of Calgary, Heritage Medical Research Building, 3300 Hospital Dr. NW, Calgary, Alberta, Canada T2N 4N1.
Shane
S
Kirby
Department of Microbiology and Infectious Diseases, Faculty of Medicine, University of Calgary, Heritage Medical Research Building, 3300 Hospital Dr. NW, Calgary, Alberta, Canada T2N 4N1.
Anthony B
AB
Schryvers
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000187:drug effects / Q000235:genetics*; D001483:Base Sequence; D002352:Carrier Proteins / Q000235:genetics*; D015964:Gene Expression Regulation, Bacterial / Q000187:drug effects / Q000235:genetics*; D006193:Haemophilus influenzae / Q000187:drug effects / Q000235:genetics*; D008969:Molecular Sequence Data; D009344:Neisseria gonorrhoeae / Q000235:genetics; D009693:Nucleic Acid Hybridization; D021382:Protein Sorting Signals / Q000235:genetics / Q000502:physiology*; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D012689:Sequence Homology, Nucleic Acid; D013211:Staphylococcus aureus / Q000235:genetics; D001618:beta-Lactamases / Q000235:genetics
D001426:Bacterial Proteins; D002352:Carrier Proteins; D021382:Protein Sorting Signals; C093280:fibrinogen-binding protein A, Staphylococcus aureus; D001618:beta-Lactamases