Table 1.
Pentapeptide insertion and site-directed mutants: a summary
| Mutant | Sub-domain | Location | Inserted sequence or point mutation(s) | Ability to substitute for Cdc1 at different expression levels | Interaction with Pol3 and Cdc27 by two-hybrid analysis (%) | ||
| Low | High | Pol3 | Cdc27 | ||||
| WT | + | + | 100 | 100 | |||
| J12 | N | 7 | Ser-GlyValProHisSer-Ile | + | + | <1 | 59 |
| E4 | N | 10 | Cys-GlyValProProCys-Glu | + | + | 16 | 62 |
| J2 | I | 25 | Tyr-ArgGlyThrProTyr-Ser | + | + | 46 | <1 |
| E2 | I | 26 | Ser-GlyValProLysTer | - | - | <1 | <1 |
| J17 | I | 28 | Gln-Ter | - | + | 2 | <1 |
| J3 | I/II | 62 | Leu-GlyValProLeuLeu-Asp | - | + | 54 | <1 |
| E1 | I/II | 66 | Ser-GlyValProGlnSer-Asp | + | + | 65 | 1 |
| J10 | II | 77 | Tyr-ArgGlyThrProTyr-Met | - | + | <1 | 30 |
| J8 | II | 83 | Lys-GlyValProLeuLys-Pro | + | + | <1 | 128 |
| E5 | II | 86 | Val-MetGlyTyrProVal-Met | + | + | 48 | 88 |
| J6 | III | 124 | Tyr-GlyValProHisTyr-Gly | - | + | <1 | <1 |
| E9 | III | 124 | Tyr-GlyGlyTyrProTyr-Gly | - | + | 24 | <1 |
| J4 | III | 127 | Ile-GlyValProArgIle-Asp | - | + | 36 | <1 |
| J1 | III | 141 | Thr-GlyValProLeuThr-Gly | - | - | <1 | <1 |
| J16 | III/IV | 160 | Val-GlyGlyThrProVal-Asp | - | - | <1 | <1 |
| J9 | III/IV | 174 | Met-ArgGlyThrProMet-Thr | + | + | 91 | 102 |
| J15 | IV/V | 203 | Gln-GlyValProLeuGln-Val | + | + | 1 | 78 |
| J22 | IV/V | 212 | Arg-GlyValProLeuArg-Gly | - | + | 25 | 7 |
| J21 | V/VI | 275 | Leu-GlyValProGlnLeu-Asp | - | + | <1 | <1 |
| A1 | VI | 292 | Pro292Ala | + | + | 39 | 44 |
| A2 | VI | 293 | Gly293Ala | + | + | <1 | 19 |
| A9 | VI | 292–293 | Pro292Ala, Gly293Ala | - | - | <1 | 58 |
| A3 | VI | 296 | Asp296Ala | - | - | <1 | 23 |
| A4 | VI | 303 | Pro303Ala | + | + | <1 | 31 |
| A5 | VII | 328 | Asn328Ala | + | + | 4 | 37 |
| A6 | VII | 329 | Pro329Ala | - | + | <1 | 6 |
| A10 | VII | 328–329 | Asn328Ala, Pro329Ala | - | - | <1 | 72 |
| A7 | VII | 344 | Gly344Ala | + | + | 9 | <1 |
| A8 | VIII | 374 | Pro374Ala | + | + | 17 | 8 |
| J7 | VIII | 393 | Met-GlyValProLeuMet-Glu | - | - | <1 | 19 |
For the pentapeptide insertion mutants, multiply isolated insertions were as follows: J2 (re-isolated as J18, J20, E10), J3 (E3, E8), J10 (J11, E6, E7), J8 (J19) and J4 (J5, J13). The location of the insertions is given as the number of the last amino acid residue N-terminal to the insertion. The insertion sequences are shown between the hyphens. Note that the target site duplication results in the amino acid immediately N-terminal to the insertion being duplicated as the fifth amino acid of the insertion (except where stop codons are inserted) and that the insertions in Cdc1-J6 and Cdc1-E9 are found at the same amino acid but are of different sequence. Quantitative two-hybrid assay values are given as percentages of the interaction seen with wild-type Cdc1 with Pol3 (in the presence of Cdm1) or Cdc27. See Results for further details.