TABLE 2.
Oligonucleotides used in this study
| Method | Primer | Sequencea | Plasmidb |
|---|---|---|---|
| PCR | PepE5 | GGAATTCTTTCCTGATGAGGTGGAGAA | |
| PepE6 | CGGGATCCAATATGGCCTGGGATTAA | ||
| BsPepE1 | GAGAATTCGGCGGGTGTGATGATGAAGCAGAT | ||
| BsPepE2 | CGGGATCCTTGATTTCGCACTCATCTTATCAT | ||
| Sequencing | PepE1 | CGTCTCTAAACGTGTTAAAT | |
| PepE3 | CAATATGGCCTGGGATTA | ||
| PepE4 | ATCCGTGTACTCGTCCCATGTT | ||
| pSE380 | AATTGTGAGCGGATAAC | ||
| Mutagenesis | PstI/XhoI | GATATCGAATTCCTCGAGCCCGGG | |
| PepE-S7A | GCTTTTATTAGCTAACTCGACGC | pCM442 | |
| PepE-S9A | GCTTTTATTAAGTAACGCGACGCTGCCGGG | pCM461 | |
| PepE-S120A | CGGCTGGGCCGCGGGCGCC | pCM437 | |
| PepE-S120C | CGGCTGGTGCGCGGGCGCC | pCM456 | |
| PepE-H19A | GCCTGGCTGGAAGCCGCTCTGCCGCTG | pCM447 | |
| PepE-H19Q | CTGGCTGGAACAAGCTCTGCCGC | pCM443 | |
| PepE-H68A | GTCACAGGGATTGCTCGTGTCGCCGATC | pCM448 | |
| PepE-H157A | GCAAATTAATCCTGCCTTCACTAACGCC | pCM439 | |
| PepE-H157Q | GCAAATTAATCCTCAATTCACTAACGCC | pCM457 | |
| PepE-H166A | CTTGCCGGAAGGTGCTAAGGGCGAGACC | pCM465 | |
| PepE-H166Q | GCCGGAAGGTCAAAAGGGCGAGAC | pCM466 | |
| PepE-D47A | CGCAAACATGGGCCGAGTACACGG | pCM450 | |
| PepE-D135A | GCACGACCAACGCTATGCCGATCG | pCM438 | |
| PepE-E172A | GAGACCCGCGCGCAGCGTATTC | pCM458 | |
| PepE-K95A | CCAGCTACTGGCAGAATCGCGTG | pCM462 | |
| PepE-K171E | GGGCGAGACCGAGGAGCAGCG | pCM453 | |
| PepE-R174A | CCGCGAGCAGGCTATTCGCGAAC | pCM454 | |
| PepE-R174E | CCGCGAGCAGGAGATTCGCGAAC | pCM455 |
a
The relevant mutagenized codon or restriction site is underlined.
b
Plasmids are derivatives of pCM389, which carries wild-type pepE.