Identification of Sphingomyelinase on the Surface of Chlamydia pneumoniae: Possible Role in the Entry into Its Host Cells
Figure 3
TLC analysis on silicic acid coated plates of the hydrolysis of the fluorescent Bdp-sphingomyelin (lane A) to Bdp-ceramide (lane D). Formation of fluorescent ceramide from Bdp-SM (50 μM final concentration) in 0.5 mM Hepes, 1.8 mM MgCl2, and 9 mM CaCl2; pH 7.4 was monitored after the addition of 3.3 × 107 (inclusion forming units (IFUs) (lane B) and 6.6 × 107 IFUs of intact C. pneumoniae (lane C) in the same buffer. Incubation for 48 hrs at 37°C was used to ensure sufficient product formation so as to allow the detection of ceramide upon TLC despite losing some of this lipid in extraction. The TLC plates were developed with 1,2-dichloroethane/methanol/water (90 : 20 : 0.5, by vol) and the lipids localized by illumination with a UV lamp.
