Table 1 Bacterial strains and plasmids used in this study
(A) Bacterial strains | ||
|---|---|---|
Names | Genotype, characteristics | Reference |
Escherichia coli | ||
JM109 | endA1, recA1, gyrA96, thi, hsdR17 (r −k , m +k ), relA1, supE44, Δ(lac-proAB), [F´ traD36, proAB, laqIqZΔM15] | Promega |
Lactococcus lactis | ||
MG1363 | plasmid free derivative of NCDO712 | Laboratory collection |
S. aureus | ||
Lowenstein (ATCC 49521) | clinical isolate, capsular polysaccharide CP5-positive | Laboratory collection |
COL | clinical isolate, methicillin resistant | Laboratory collection |
htrA 1 | COL htrA 1 :: cat, CmR | [57] |
htrA 2 | COL htrA 2 :: spc, SpcR | [57] |
(B) Plasmids | ||
|---|---|---|
Names | Characteristics | Reference |
pGEMT | T-tailed PCR product Cloning vector, AmpR | Promega |
pVE8124 | pGEMT derivative where recombinant htrA 1r ORF is cloned htrA 1r encodes HtrA1-ΔTM-Ser255Ala-His6 protein | This work |
pVE8125 | pGEMT derivative where recombinant htrA 2r ORF is cloned htrA 2r encodes SPExp4-HtrA2-ΔTM-Ser619Ala-His6 protein | This work |
pLB145 | pWV01; CmR; expression-secretion vector where exp4 SP -nuc is cloned under the control of a lactococcal expression system, P zit zitR exp4 SP -nuc encodes a hybrid protein between a lactococcal signal peptide (SPExp4) and the mature secreted form of the staphylococcal nuclease (NucB) | [4] |
pVE8126 | pLB145 derivative where htrA 1r ORF is cloned in place of nuc ORF to be fused in frame to exp4 SP exp4 SP -htrA 1r encodes a hybrid precursor leading to secreted rHtrA1 protein | This work |
pVE8127 | pLB145 derivative where htrA 2r ORF is cloned in place of exp4 SP -nuc ORF htrA 2r encodes a hybrid precursor leading to secreted rHtrA2 protein | This work |