Lipopolysaccharide neutralizing peptide-porphyrin conjugates for effective photoinactivation and intracellular imaging of gram-negative bacteria strains.
ABSTRACT A simple and specific strategy based on the bioconjugation of a photosensitizer protophophyrin IX (PpIX) with a lipopolysaccharide (LPS) binding antimicrobial peptide YI13WF (YVLWKRKRKFCFI-Amide) has been developed for the effective fluorescent imaging and photodynamic inactivation of Gram-negative bacterial strains. The intracellular fluorescent imaging and photodynamic antimicrobial chemotherapy (PACT) studies supported our hypothesis that the PpIX-YI13WF conjugates could serve as efficient probes to image the bacterial strains and meanwhile indicated the potent activities against Gram-negative bacterial pathogens especially for those with antibiotics resistance when exposed to the white light irradiation. Compared to the monomeric PpIX-YI13WF conjugate, the dimeric conjugate indicated the stronger fluorescent imaging signals and higher photoinactivation toward the Gram-negative bacterial pathogens throughout the whole concentration range. In addition, the photodynamic bacterial inactivation also demonstrated more potent activity than the minimum inhibitory concentration (MIC) values of dimeric PpIX-YI13WF conjugate itself observed for E. coli DH5a (~4 times), S. enterica (~8 times), and other Gram-negative strains including antibiotic-resistant E. coli BL21 (~8 times) and K. pneumoniae (~16 times). Moreover, both fluorescent imaging and photoinactivation measurements also demonstrated that the dimeric PpIX-YI13WF conjugate could selectively recognize bacterial strains over mammalian cells and generate less photo damage to mammalian cells. We believed that the enhanced fluorescence and bacterial inactivation were probably attributed to the higher binding affinity between dimeric photosensitizer peptide conjugate and LPS components on the surface of bacterial strains, which were the results of efficient multivalent interactions.
Article: Photoinactivation of Gram positive and Gram negative bacteria with the antimicrobial peptide (KLAKLAK)(2) conjugated to the hydrophilic photosensitizer eosin Y.[show abstract] [hide abstract]
ABSTRACT: We test the hypothesis that the antimicrobial peptide (KLAKLAK)(2), enhances the photodynamic activity of the photosensitizer eosin Y upon conjugation. The conjugate eosin-(KLAKLAK)(2) was obtained by solid-phase peptide synthesis. Photoinactivation assays were performed against the Gram-negative bacteria Escherichia coli, Pseudomonas aeruginosa, and multi-drug resistant Acinetobacter baumannii AYE, as well as the Gram positive bacteria Staphylococcus aureus, and Staphylococcus epidermidis. Partitioning assays were performed with E. coli and S. aureus. Photohemolysis and photo-killing assays were also performed to assess the photodynamic activity of the conjugate towards mammalian cells. Eosin-(KLAKLAK)(2) photo-inactivates 99.999% of 10(8) CFU/mL of most bacteria tested at a concentration of 1 μM or below. In contrast, neither eosin Y nor (KLAKLAK)(2) cause any significant photoinactivation under similar conditions. The increase in photodynamic activity of the photosensitizer conferred by the antimicrobial peptide is in part due to the fact that (KLAKLAK)(2) promotes the association of eosin Y to bacteria. Eosin-(KLAKLAK)(2) does not significantly associate with red blood cells or the cultured mammalian cell lines HaCaT, COS-7 and COLO 316. Consequently, little photo-damage or photo-killing is observed with these cells under conditions for which bacterial photoinactivation is achieved. The peptide (KLAKLAK)(2) therefore significantly enhances the photodynamic activity of eosin Y towards both Gram-positive and Gram-negative bacteria while interacting minimally with human cells. Overall, our results suggest that antimicrobial peptides such as (KLAKLAK)(2) might serve as attractive agents that can target photosensitizers to bacteria specifically.Bioconjugate Chemistry 12/2012; · 4.93 Impact Factor