Details

Autor(en) / Beteiligte

• Roten, C.A.H.
• Pagni, M.
• Margot, P.
• Touri, F.
• Karamata, D.

Titel

Specific Labeling of Diaminopimelate: A Radioassay for the Determination of the Peptidoglycan Cross-Linking Index

Ist Teil von

• Analytical biochemistry, 1994-12, Vol.223 (2), p.208-211

Ort / Verlag

United States: Elsevier Inc

Erscheinungsjahr

1994

Quelle

Elsevier Journal Backfiles on ScienceDirect (DFG Nationallizenzen)

Beschreibungen/Notizen

• A radioassay for the determination of the peptidoglycan cross-linking index (CLI) was devised. It is based on specific radioactive labeling of diaminopimelic acid (DAP) by diverting [14C]aspartate into the DAP pathway, while inhibiting incorporation of label into other cell wall components. Purified [14C]DAP-labeled cell walls were treated with fluorodinitrobenzene, hydrolyzed, and chromatographed by TLC. The radioactivity in well-separated mono dinitrophenyl-diaminopimelate (DNP-DAP) and DAP spots was counted and the CLI was determined from the ratio of DAP to the total of mono DNP-DAP and DAP counts. The method, suitable for bacteria such as Bacillus subtilis unable to incorporate exogenous DAP, can be applied to other systems. A CLI of 50.8 ± 1.3% and 55.5 ± 0.9% was obtained for B. subtilis 168 cells growing exponentially in rich and minimal medium, respectively. Comparison of these to results previously obtained on B. subtilis suggested the existence of a hitherto unreported peptidoglycan endopeptidase activity.