TABLE 1.
Types and frequencies of etnE mutations in JS623 cultures during VC adaptation
| Culture | Ethene exposure timea,c (days) | VC exposure timeb,c (days) | Predicted EtnE mutant type | No. of clones |
|---|---|---|---|---|
| Wild-type | 21 | 0 | None | 15 |
| JS623 | A60Ad | 1 | ||
| N98I | 1 | |||
| K286R | 1 | |||
| F297S | 1 | |||
| T311T | 1 | |||
| 100 | 0 | None | 13 | |
| V53A, R128L, V304A | 1 | |||
| E210G, W293R | 1 | |||
| H123H, N343N | 1 | |||
| L273S | 1 | |||
| S138N, A258V | 2 | |||
| T268T | 1 | |||
| C214S, A253V | 1 | |||
| JS623-E | 15 | 97 | W243G | 8 |
| R257L | 5 | |||
| V244G | 2 | |||
| W243R | 1 | |||
| W243L, L245V | 1 | |||
| L245P | 1 | |||
| 15 | 178 | W243G | 12 | |
| R257L | 7 | |||
| W243R | 1 | |||
| W243G, R257L | 1 | |||
| JS623-T | 15 | 86 | R257L | 8 |
| R257L, E179G | 1 | |||
| R257L, I165I | 1 | |||
| R257L, E160G, D303G | 1 | |||
| W243R | 1 | |||
| W243R, D68N | 1 | |||
| W243R, S80P | 1 | |||
| W243R, D317Y | 1 | |||
| W243R, F175F, I385V | 1 | |||
| V244G | 1 | |||
| V244G, G223E | 1 | |||
| V244G, N369D | 1 | |||
| V244G, T134T, H352R | 1 | |||
| 15 | 235 | R257L | 10 | |
| R257L, F47S | 1 | |||
| R257L, I82V | 1 | |||
| R257L, T130A | 1 | |||
| R257L, A166T | 1 | |||
| R257L, W293C | 1 | |||
| R257L, N343S | 1 | |||
| R257L, S347S | 1 | |||
| R257L, F37F, L238G | 1 | |||
| R257L, N98S, L127L, F262S | 1 | |||
| 15 | 289 | R257L | 10 | |
| R257L, D25N | 1 | |||
| R257L, D45G | 1 | |||
| R257L, Q58*stop | 1 | |||
| R257L, A79A | 1 | |||
| R257L, I167T | 1 | |||
| R257L, N274S | 1 | |||
| R257L, L127L,H219Y | 1 | |||
| R257L, V218V, S334P | 1 | |||
| R257L, N98D, A166A, T302I | 1 |
Consecutive culturing time on ethene since the strain was received in the Mattes laboratory. VC-adapted variants were cultured for 15 days on ethene before transfer to VC, while the wild-type strain was grown only on ethene and was sampled at 21 and 100 days.
Consecutive culturing time on VC since transfer from ethene (JS623-E) or TSAG (JS623-T) medium. Zero indicates that the wild-type JS623 cultures were not exposed to VC.
Cultures were routinely archived at −80°C after reaching an OD600 of ∼0.3 and were analyzed later for the presence of missense mutations. This experimental design did not allow for the calculation of generation times.
Amino acid location in the EtnE protein sequence (GenBank accession no. FJ602754).