TABLE 1.
Bacterial strains, plasmids, and oligonucleotides used in this study
| Strain, plasmid, or nucleotide | Relevant characteristic(s) or sequencea | Source or reference |
|---|---|---|
| E. coli JM109 (SN3688) | Plasmid storage and expression of lcp | |
| E. coli XL1-Blue (SN4827) | Transformation strain | Stratagene |
| pUC9::strep-lcpK30 (SN5339) | Cloning vector for lcpK30, Apr | |
| p4782.1 (SN3513) | Mobilizable broad host range expression vector, Kmr | 21 |
| p4782.1::strep-lcpK30 (SN5496) | Coding sequence of strep-lcpK30 under rhamnose promoter control, Kmr | 12 |
| p4782.1::strep-lcpRr (SN5760) | Coding sequence of strep-lcpRr under rhamnose promoter control, Kmr | 15 |
| p4782.1::lcpK30 R164A (SN6034) | R164A mutein of SN5496 | 12 |
| p4782.1::lcpK30 T168A (SN6039) | T168A mutein of SN5496 | 12 |
| p4782.1::lcpK30 H198A (SN5528) | H198A mutein of SN5496 | 12 |
| p4782.1::lcpK30 H203A (SN5530) | H203A mutein of SN5496 | 12 |
| p4782.1::lcpK30 H232A (SN6035) | H232A mutein of SN5496 | 12 |
| p4782.1::lcpK30 H259A (SN6040) | H259A mutein of SN5496 | 12 |
| p4782.1::lcpK30 H266A (SN6041) | H266A mutein of SN5496 | 12 |
| p4782.1::lcpK30 R195A (SN5527) | R195A mutein of SN5496 | 12 |
| p4782.1::lcpK30 R202A (SN5529) | R202A mutein of SN5496 | 12 |
| p4782.1::lcpK30 R328A (SN6042) | R328A mutein of SN5496 | 12 |
| Fw-R164A | GACATGAAGGACGCAATCGCTAAGACCGC | |
| Rev-R164A | GCGGTCTTAGCGATTGCGTCCTTCATGTC | |
| Fw-T168A | ACCGAATCGCTAAGGCCGCACGTCTCGGT | |
| Rev-T168A | ACCGAGACGTGCGGCCTTAGCGATTCGGT | |
| Fw-H198A | ACGCATGGTGGCAGCGGCTGTTCGCC | |
| Rev-H198A | GAACAGCCGCTGCCACCATGCGTGTC | |
| Fw-H203A | GGCTGTTCGCGCACTGCTGCCGCAGT | |
| Rev-H203A | GCGGCAGCAGTGCGCGAACAGCCGCG | |
| Fw-H232A | ATGGTCACCTGGGCAAGCCTGGCCACT | |
| Rev-H232A | AGTGGCCAGGCTTGCCCAGGTGACCAT | |
| Fw-H259A | GCGGAGGCTTACCTTGCAGTGTGGCAGGT | |
| Rev-H259A | GACCTGCCACACTGCAAGGTAAGCCTCCG | |
| Fw-H266A | TGGCAGGTCAGTGCGGCAATGCTCGGAGT | |
| Rev-H266A | GACTCCGAGCATTGCCGCACTGACCTGCC | |
| Fw-R195A | AGTGAAGACAGCAATGGTGCACGCGG | |
| Rev-R195A | CGTGCACCATTGCTGTCTTCACTGCG | |
| Fw-R202A | CGCGGCTGTTGCACACCTGCTGCCGC | |
| Rev-R202A | GCAGCAGGTGTGCAACAGCCGCGTGC | |
| Fw-R328A | GGCGCGTTCTCGGCGTACACACTCGGCGG | |
| Rev-R328A | GCCGCCGAGTGTGTACGCCGAGAACGCGC | |
| pInt_F | CCCATTTTCCTGTCAGTAAC | |
| pInt_R | CTCCACGGGGAGAGCCTGAG |
a
Kmr, kanamycin resistance; Apr, ampicillin resistance.