title: Detection of Listeria monocytogenes with short peptide fragments from class IIa bacteriocins as recognition elements.
abstract: We employed a direct peptide-bacteria binding assay to screen peptide fragments for high and specific binding to Listeria monocytogenes. Peptides were screened from a peptide array library synthesized on cellulose membrane. Twenty four peptide fragments (each a 14-mer) were derived from three potent anti-listerial peptides, Leucocin A, Pediocin PA1, and Curvacin A, that belong to class IIa bacteriocins. Fragment Leu10 (GEAFSAGVHRLANG), derived from the C-terminal region of Leucocin A, displayed the highest binding among all of the library fragments toward several pathogenic Gram-positive bacteria, including L. monocytogenes, Enterococcus faecalis, and Staphylococcus aureus. The specific binding of Leu10 to L. monocytogenes was further validated using microcantilever (MCL) experiments. Microcantilevers coated with gold were functionalized with peptides by chemical conjugation using a cysteamine linker to yield a peptide density of ∼4.8×10(-3) μmol/cm2 for different peptide fragments. Leu10 (14-mer) functionalized MCL was able to detect Listeria with same sensitivity as that of Leucocin A (37-mer) functionalized MCL, validating the use of short peptide fragments in bacterial detection platforms. Fragment Leu10 folded into a helical conformation in solution, like that of native Leucocin A, suggesting that both Leu10 and Leucocin A may employ a similar mechanism for binding target bacteria. The results show that peptide-conjugated microcantilevers can function as highly sensitive platforms for Listeria detection and hold potential to be developed as biosensors for pathogenic bacteria.
authors.affiliation: Faculty of Pharmacy and Pharmaceutical Sciences, University of Alberta , Edmonton, Alberta T6G 2E1, Canada.
authors.forename: Sarfuddin
authors.initials: S
authors.lastname: Azmi
authors.forename: Keren
authors.initials: K
authors.lastname: Jiang
authors.forename: Michael
authors.initials: M
authors.lastname: Stiles
authors.forename: Thomas
authors.initials: T
authors.lastname: Thundat
authors.forename: Kamaljit
authors.initials: K
authors.lastname: Kaur
mesh_terms: D001430:Bacteriocins / Q000032:analysis / Q000737:chemistry*; D015374:Biosensing Techniques / Q000379:methods*; D008089:Listeria monocytogenes / Q000737:chemistry* / Q000302:isolation & purification*; D010446:Peptide Fragments / Q000032:analysis / Q000737:chemistry*
chemical_list: D001430:Bacteriocins; D010446:Peptide Fragments
keywords: Listeria monocytogenes; antimicrobial peptide; bacterial detection; class IIa bacteriocins; microcantilever; peptide array