Adhesion characteristics of Listeria adhesion protein (LAP)-expressing Escherichia coli to Caco-2 cells and of recombinant LAP to eukaryotic receptor Hsp60 as examined in a surface plasmon resonance sensor

• Published in: FEMS microbiology letters 2006-03, Vol.256 (2), p.324-332
• Main Authors: Kim, Kwang-Pyo, Jagadeesan, Balamurugan, Burkholder, Kristin M., Jaradat, Ziad W., Wampler, Jennifer L., Lathrop, Amanda A., Morgan, Mark T., Bhunia, Arun K.
• Format: Article
• Language: English
• Subjects: Acetaldehyde; Acetaldehyde dehydrogenase; Adhesins, Bacterial - biosynthesis; Adhesins, Bacterial - genetics; Adhesins, Bacterial - metabolism; Adhesion; adhesion kinetics; Alcohol; alcohol acetaldehyde dehydrogenase; Alcohol Dehydrogenase - biosynthesis; Alcohol Dehydrogenase - genetics; Alcohol Dehydrogenase - metabolism; Antibodies; Avidity; Bacteria; Bacterial Adhesion; Bacteriology; Binding; Biological and medical sciences; Caco-2 Cells; Chaperonin 60 - metabolism; Dehydrogenase; Dehydrogenases; E coli; Escherichia coli; Escherichia coli - genetics; Escherichia coli - physiology; Fundamental and applied biological sciences. Psychology; Genetics; Heat shock proteins; Hsp60; Hsp60 protein; Humans; Internalin; Kinetics; Listeria; Listeria adhesion protein; Listeria monocytogenes; Listeria monocytogenes - enzymology; Listeria monocytogenes - genetics; Metabolism. Enzymes; Microbiology; Permeability, membrane transport, intracellular transport; Protein Binding; Proteins; Recombinant Proteins - biosynthesis; Recombinant Proteins - genetics; Recombinant Proteins - metabolism; Resonance; SPR; Surface Plasmon Resonance
• ISSN: 0378-1097; 1574-6968
• DOI: 10.1111/j.1574-6968.2006.00140.x

Abstract Listeria adhesion protein (LAP) is an important adhesion factor in Listeria monocytogenes and interacts with its cognate receptor, mammalian heat shock protein 60 (Hsp60). The genetic identity of LAP was determined to be alcohol acetaldehyde dehydrogenase (Aad). A recombinant Escherichia coli strain expressing aad confirmed the involvement of Aad in adhesion to Caco-2 cells. Binding kinetics (ka) of recombinant LAP (rLAP) to Hsp60 was examined in a surface plasmon resonance sensor and was determined to be 5.35 × 108 M−1 s−1 and it was equivalent to the binding of anti-Hsp60 antibody (ka=2.15 × 109 M−1 s−1) to Hsp60. In contrast, Internalin B, an adhesion/invasion protein from L. monocytogenes, used as a control, had binding kinetics (ka) of only 2.9 × 106 M−1 s−1. The KD value of rLAP was 1.68 × 10−8 M, which was significantly lower than Internalin B (KD=6.5 × 10−4 M). These results suggest that Hsp60 has significantly higher avidity for anti-Hsp60 antibody and LAP than Internalin B. In summary, LAP is identified as an alcohol acetaldehyde dehydrogenase and binding of recombinant E. coli to Caco-2 cells or rLAP to Hsp60 protein was found to be highly specific.