title: Gamma-phage lysin PlyG sequence-based synthetic peptides coupled with Qdot-nanocrystals are useful for developing detection methods for Bacillus anthracis by using its surrogates, B. anthracis-Sterne and B. cereus-4342.
abstract: BACKGROUND
  Previous reports of site-directed deletion analysis on gamma (gamma)-phage lysin protein (PlyG) have demonstrated that removal of a short amino acid sequence in the C-terminal region encompassing a 10-amino acid motif (190LKMTADFILQ199) abrogates its binding activity specific to the cell wall of Bacillus anthracis. Whether short synthetic peptides representing the10-amino acid PlyG putative binding motif flanked by surrounding N- and C-terminal residues also selectively bind to the bacterial cell wall has not been evaluated. If such peptides do demonstrate selective binding to the cell wall, they could serve as bio-probes towards developing detection technologies for B. anthracis. Furthermore, by using B. anthracis (Sterne, 34F2), an animal vaccine and B. cereus-4342, a gamma-phage susceptible rare strain as surrogates of B. anthracis, development of proof-of-concepts for B. anthracis are feasible.
  
  
  RESULTS
  Using four different methods, we evaluated six synthetic peptides representing the putative binding motif including flanking sequences (PlyG-P1 through P6) for the bacterial cell wall binding capacity. Our analysis identified PlyG-P1, PlyG-P3 and PlyG-P5 to have binding capability to both B. anthracis (Sterne, 34F2) and B. cereus-4342. The peptides however did not bind to B. cereus-11778, B. thuringiensis, and B. cereus-10876 suggesting their specificity for B. anthracis-Sterne and B. cereus-4342. PlyG-P3 in combination with fluorescent light microscopy detected even a single bacterium in plasma spiked with the bacteria.
  
  
  CONCLUSIONS
  Overall, these studies illustrate that the short 10-amino acid sequence 'LKMTADFILQ' in fact is a stand-alone bacterial cell wall-binding motif of PlyG. In principle, synthetic peptides PlyG-P1, PlyG-P3 and PlyG-P5, especially PlyG-P3 coupled with Qdot-nanocrystals are useful as high-sensitivity bio-probes in developing detection technologies for B. anthracis.
authors.affiliation: Section of Cell biology, Laboratory of Cellular Hematology, Center for Biologics Evaluation and Research, FDA, Bethesda, MD 20892, USA.
authors.forename: Shilpakala
authors.initials: S
authors.lastname: Sainathrao
authors.forename: Ketha V Krishna
authors.initials: KV
authors.lastname: Mohan
authors.forename: Chintamani
authors.initials: C
authors.lastname: Atreya
mesh_terms: D000595:Amino Acid Sequence; D000881:Anthrax / Q000175:diagnosis / Q000382:microbiology; D017103:Bacillus Phages; D001408:Bacillus anthracis / Q000302:isolation & purification*; D001665:Binding Sites; D002473:Cell Wall / Q000382:microbiology; D004797:Enzyme-Linked Immunosorbent Assay; D008969:Molecular Sequence Data; D009238:N-Acetylmuramoyl-L-alanine Amidase / Q000378:metabolism*; D010455:Peptides / Q000138:chemical synthesis / Q000378:metabolism*; D045663:Quantum Dots; D012680:Sensitivity and Specificity; D014764:Viral Proteins / Q000378:metabolism*
chemical_list: D010455:Peptides; D014764:Viral Proteins; C465450:PlyG lysin, gamma phage; D009238:N-Acetylmuramoyl-L-alanine Amidase