Incorporation of a Valine-Leucine-Lysine-Containing Substrate in the Bacterial Cell Wall.
The emergence of antibiotic-resistant bacteria is a major public health threat, and therefore novel antimicrobial targets and strategies are urgently needed. In this regard, cell-wall-associated proteases are envisaged as interesting antimicrobial targets due to their role in cell wall remodeling. Here, we describe the discovery and characteristics of a protease substrate that is processed by a bacterial cell-wall-associated protease. Stationary-phase grown Gram-positive bacteria were incubated with fluorogenic protease substrates, and their cleavage and covalent incorporation into the cell wall was analyzed. Of all of the substrates used, only one substrate, containing a valine-leucine-lysine (VLK) motif, was covalently incorporated into the bacterial cell wall. Linkage of the VLK-peptide substrate appeared unrelated to sortase A and B activity, as both wild-type and sortase A and B knock out Staphylococcus aureus strains incorporated this substrate into their cell wall with comparable efficiency. Additionally, the VLK-peptide substrate showed significantly higher incorporation in the cell wall of VanA-positive Enterococcus faecium strains than in VanB- and vancomycin-susceptible isolates. In conclusion, the VLK-peptide substrate identified in this study shows promise as a vehicle for targeting antimicrobial compounds and diagnostic contrast agents to the bacterial cell wall.
Department of Medical Microbiology and Infectious Diseases, Erasmus MC , Wytemaweg 80, 3015 CE Rotterdam, The Netherlands.|Department of Oral Biochemistry, Academic Centre for Dentistry Amsterdam, University of Amsterdam and VU University Amsterdam , Gustav Mahlerlaan 3004, 1081 LA Amsterdam, The Netherlands.
Silvie
S
Hansenová Maňásková
Department of Oral Biochemistry, Academic Centre for Dentistry Amsterdam, University of Amsterdam and VU University Amsterdam , Gustav Mahlerlaan 3004, 1081 LA Amsterdam, The Netherlands.
Floris J
FJ
Bikker
Department of Oral Biochemistry, Academic Centre for Dentistry Amsterdam, University of Amsterdam and VU University Amsterdam , Gustav Mahlerlaan 3004, 1081 LA Amsterdam, The Netherlands.
Kamran
K
Nazmi
Department of Medical Microbiology and Infectious Diseases, Erasmus MC , Wytemaweg 80, 3015 CE Rotterdam, The Netherlands.
Rianne
R
van Zuidam
Optical Imaging Center, Department of Pathology, Josephine Nefkens Institute, Erasmus MC , Dr Molewaterplein 50, Rotterdam 3015 GE, The Netherlands.
Johan A
JA
Slotman
Optical Imaging Center, Department of Pathology, Josephine Nefkens Institute, Erasmus MC , Dr Molewaterplein 50, Rotterdam 3015 GE, The Netherlands.
Wiggert A
WA
van Cappellen
Optical Imaging Center, Department of Pathology, Josephine Nefkens Institute, Erasmus MC , Dr Molewaterplein 50, Rotterdam 3015 GE, The Netherlands.
Adriaan B
AB
Houtsmuller
Department of Oral Biochemistry, Academic Centre for Dentistry Amsterdam, University of Amsterdam and VU University Amsterdam , Gustav Mahlerlaan 3004, 1081 LA Amsterdam, The Netherlands.
Enno C I
EC
Veerman
Department of Medical Microbiology and Infectious Diseases, Erasmus MC , Wytemaweg 80, 3015 CE Rotterdam, The Netherlands.|Department of Oral Biochemistry, Academic Centre for Dentistry Amsterdam, University of Amsterdam and VU University Amsterdam , Gustav Mahlerlaan 3004, 1081 LA Amsterdam, The Netherlands.
Wendy E
WE
Kaman
D020816:Amino Acid Motifs; D019881:Aminoacyltransferases / Q000235:genetics / Q000378:metabolism; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D019729:Carbon-Oxygen Ligases / Q000378:metabolism; D002473:Cell Wall / Q000737:chemistry* / Q000378:metabolism; D003546:Cysteine Endopeptidases / Q000235:genetics / Q000378:metabolism; D016984:Enterococcus faecium / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D006094:Gram-Positive Bacteria / Q000166:cytology* / Q000378:metabolism; D007930:Leucine / Q000737:chemistry; D008239:Lysine / Q000737:chemistry; D008826:Microbial Sensitivity Tests; D010455:Peptides / Q000493:pharmacokinetics*; D013211:Staphylococcus aureus / Q000166:cytology / Q000235:genetics; D014633:Valine / Q000737:chemistry
D001426:Bacterial Proteins; D010455:Peptides; C071437:VanA ligase, Bacteria; C509821:sortase B; D019881:Aminoacyltransferases; C120542:sortase A; D003546:Cysteine Endopeptidases; D019729:Carbon-Oxygen Ligases; D007930:Leucine; D014633:Valine; D008239:Lysine