EscU auto-cleavage results in a 10 kDa C-terminal product that is membrane associated in EPEC. (A) Isolated membrane fractions were probed with anti-HA and anti-FLAG antibodies to assess EscU auto-cleavage status. Membrane localization of EscJ is unchanged in escU null mutants (lane 2) and therefore this protein served as an internal control for the individual membrane fractions. The approximate 10 kDa C-terminal EscU auto-cleavage product (detected with anti-FLAG antibodies) along with the 29 kDa HA-tagged N-terminal product (detected with anti-HA antibodies) both partitioned to the membrane fraction (denoted by arrows). Uncleaved EscU is also membrane associated and appeared as a 39 kDa species. (B) The same membrane fractions were probed with anti-EscN antibodies to detect membrane associated EscN levels. A ΔescN mutant membrane preparation was included to demonstrate the specificity of the antibody.
Thomassin et al. BMC Microbiology 2011 11:205 doi:10.1186/1471-2180-11-205