Document Detail


Intracellular toxicity of proline-rich antimicrobial peptides shuttled into mammalian cells by the cell-penetrating peptide penetratin.
MedLine Citation:
PMID:  22850523     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
The health threat caused by multiresistant bacteria has continuously increased and recently peaked with pathogens resistant to all current drugs. This has triggered intense research efforts to develop novel compounds to overcome the resistance mechanisms. Thus, antimicrobial peptides (AMPs) have been intensively studied, especially the family of proline-rich AMPs (PrAMPs) that was successfully tested very recently in murine infection models. PrAMPs enter bacteria and inhibit chaperone DnaK. Here, we studied the toxicity of intracellular PrAMPs in HeLa and SH-SY5Y cells. As PrAMPs cannot enter most mammalian cells, we coupled the PrAMPs with penetratin (residues 43 to 58 in the antennapedia homeodomain) via a C-terminally added cysteine utilizing a thioether bridge. The resulting construct could transport the covalently linked PrAMP into mammalian cells. Penetratin ligation reduced the MIC for Gram-negative Escherichia coli only slightly (1 to 8 μmol/liter) but increased the activity against the Gram-positive Micrococcus luteus up to 32-fold (MIC ≈ 1 μmol/liter), most likely due to more effective penetration through the bacterial membrane. In contrast to native PrAMPs, the penetratin-PrAMP constructs entered the mammalian cells, aligned around the nucleus, and associated with the Golgi apparatus. At higher concentrations, the constructs reduced the cell viability (50% inhibitory concentration [IC(50)] ≈ 40 μmol/liter) and changed the morphology of the cells. No toxic effects or morphological changes were observed at concentrations of 10 μmol/liter or below. Thus, the IC(50) values were around 5 to 40 times higher than the MIC values. In conclusion, PrAMPs are in general not toxic to mammalian cells, as they do not pass through the membrane. When shuttled into mammalian cells, however, PrAMPs are only slightly cross-reactive to mammalian chaperones or other intracellular mammalian proteins, providing a second layer of safety for in vivo applications, even if they can enter some human cells.
Authors:
Anne Hansen; Ingo Schäfer; Daniel Knappe; Peter Seibel; Ralf Hoffmann
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't     Date:  2012-07-30
Journal Detail:
Title:  Antimicrobial agents and chemotherapy     Volume:  56     ISSN:  1098-6596     ISO Abbreviation:  Antimicrob. Agents Chemother.     Publication Date:  2012 Oct 
Date Detail:
Created Date:  2012-09-13     Completed Date:  2013-02-01     Revised Date:  2013-07-12    
Medline Journal Info:
Nlm Unique ID:  0315061     Medline TA:  Antimicrob Agents Chemother     Country:  United States    
Other Details:
Languages:  eng     Pagination:  5194-201     Citation Subset:  IM    
Affiliation:
Institute of Bioanalytical Chemistry, Faculty of Chemistry and Mineralogy, Universität Leipzig, Leipzig, Germany.
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MeSH Terms
Descriptor/Qualifier:
Animals
Anti-Infective Agents / adverse effects*,  chemical synthesis,  chemistry
Carrier Proteins / chemistry*
Cell Line
Cell-Penetrating Peptides / chemistry*
Escherichia coli / drug effects
HeLa Cells
Humans
Inhibitory Concentration 50
Mice
Microbial Sensitivity Tests
Micrococcus luteus / drug effects
Chemical
Reg. No./Substance:
0/Anti-Infective Agents; 0/Carrier Proteins; 0/Cell-Penetrating Peptides; 0/penetratin
Comments/Corrections

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