Table 2.
Insertions of the eight-codon Strep tag at various positions into lcrV and its effect on Yersinia pestis type III secretion of YopE or LcrV
| Plasmid | % type III secretion ([S]/[S+P])a |
|||
|---|---|---|---|---|
| KIM D27 (wild type) |
KLD29 (ΔlcrV) |
|||
| YopE | LcrV | YopE | LcrV | |
| None | 47.1 | 21.3 | 3.7 | NDb |
| plcrVW22703 | 50.3 | 37.8 | 51.1 | 31.8 |
| plcrVS1 | 52.3 | 25.8 | 60.8 | 24.1 |
| plcrVS20 | 4.6 | 16.8 | 7.7 | 22.7 |
| plcrVS55 | 4.1 | 15.8 | 13.2 | 21.9 |
| plcrVS170 | 48.4 | 39.7 | 45.6 | 36.3 |
| plcrVS228 | 49.2 | 38.9 | 63.4 | 37.6 |
| plcrVS270 | 54.1 | 38.5 | 2.8 | 1.1 |
Type III secretion of Y. pestis strains without or with lcrV plasmids was measured after 3 h of growth in cultures that were grown in media with chelated calcium ions at 37°C. Following centrifugation, the extracellular medium was separated into the supernatant (S) and the bacterial pellet (P), and proteins in both fractions were precipitated with trichloroacetic acid and analyzed by immunoblotting with specific antibodies against the type III secretion substrates YopE and LcrV. Secretion was quantified by calculating the percent amount of secreted protein [S] compared to protein in both the medium and the pellet [S+P].
ND, no immunoreactive signal detected.