Table 1.
Sequence variation observed in bla variants derived from the BLAΔ175 library and their influence on TEM-1 activity towards Amp in vivo
| Amino acid substitutionsa | Nucleotide substitutionsb | Base changesc | Amp MIC (μg/ml)d |
|---|---|---|---|
| Wild type | − | − | 4096 |
| M1N-S2G | atgagt→aatggt | 3 | NG |
| R9V | cgt→gtt | 2 | 512 |
| D38E-Q39P | gatcag→gaaccg | 2 | 256 |
| L40ter | ttgggt→tgatgt | 3 | NG |
| G41S | ggt→agt | 1 | 4096 |
| N52K-S53A | aacagc→aaggcc | 3 | 1024 |
| M69R-S70C | atgagc→aggtgc | 2 | NG |
| L76L C77R | ctatgt→ctacgt | 1 | NG |
| C77P | tgt→cca | 3 | NG |
| A79V | gcg→gta | 2 | 2048 |
| V103V E104G | gttgag→gtcggg | 2 | 2048 |
| V103V E104L | gttgag→gtgctg | 3 | 1024 |
| S106T | tca→acc | 2 | 256 |
| S106C | tca→tgt | 2 | 256 |
| P107A | cca→gct | 2 | 256 |
| T114T D115Y | acggat→acctat | 2 | NG |
| M117I | atg→ata | 1 | 1024 |
| T118M-V119I | acagta→atgata | 3 | 512 |
| R120R | aga→agg | 1 | 4096 |
| C123ter | tgcagt→tgatgt | 2 | NG |
| A125G-A126S | gctgcc→ggatcc | 3 | 64 |
| T128T M129G | accatg→actggg | 3 | 512 |
| A135D-N136Y | gccaac→gattac | 3 | NG |
| M155S-G156W | atgggg→agttgg | 3 | 64 |
| M155I-G156Q | atgggg→attcag | 3 | 1024 |
| M155I-G156E | atgggg→atcgag | 2 | 1024 |
| P167P E168L | ccggag→cctttg | 3 | 1024 |
| T181D | acg→gac | 3 | NG |
| R191R K192ter | cgcaaa→cgttaa | 2 | NG |
| L193L-L194L | ctatta→ctccta | 2 | 4096 |
| L194C-T195P | ttaact→tgtcct | 3 | NG |
| R204R Q205R | cggcaa→cgccga | 2 | 4096 |
| W210 fs | gac tgg atg | – | NG |
| →ga––ac atg | |||
| A213G | gcg→ggt | 2 | 4096 |
| P226P A227T | ccggct→ccaact | 2 | 4096 |
| S235 fs | tct gga→tcg ––a | – | NG |
| L250H-G251R | ctgggg→cacagg | 3 | NG |
| L250L G251R | ctgggg→ctacgg | 2 | NG |
| R259I | cgt→atc | 3 | 256 |
| T266K-G267S | acgggc→aagagc | 2 | 512 |
| Q269L | cag→ctg | 1 | 4096 |
| M272R | atg→agg | 1 | 512 |
| D273D E274Q | gatgaa→gaccaa | 2 | 2048 |
| E274P | gaa→cca | 2 | 64 |
| R277R Q278W | agacag→aggtgg | 3 | 4096 |
| Q278C | cag→tgt | 3 | 2048 |
| A280A E281S | gctgag→gcgtcg | 3 | 2048 |
| I282T | ata→acc | 2 | 64 |
| K288N-H289V | aagcat→aacgtt | 3 | 512 |
aSubscripted sequences represent silent nucleotide changes to the codon. fs represent frameshift. Residues numbered according to standard system (25).
bBold and underlined sequences represent the known nucleotides deleted from bla and replaced by SubSeqNNN.
cActual number of nucleotide differences between the variant and wild-type bla.
dNG, no cell growth observed on 32 μg/ml Amp LB agar upon removal of SubSeq and reconstitution of the full length bla.