hZW10 N-terminal point mutants analyzed for kinetochore localization and hZwint-1 interaction
| hZW10 point mutationa . | Kinetochore localization . | hZwint-1 yeast two-hybrid interaction . | β-Galactosidase assay (± one SD)b . |
|---|---|---|---|
| HS11AA | Positive | Positive | ND |
| GE36AA | Positive | Positive | ND |
| S47A | Positive | Positive | ND |
| EF48AA | Positive | Positive | ND |
| S52A | Positive | Positive | ND |
| S55A | Positive | Positive | 449 ± 139 |
| GLI58AAA | Positive | Negative | 2.7 ± 0.8 |
| SE67AA | Positive | Negative | 1.8 ± 0.9 |
| DI69AA | Positive | Negative | 1.2 ± 0.4 |
| SE79AA | Positive | Positive | 131 ± 41 |
| hZW10 point mutationa . | Kinetochore localization . | hZwint-1 yeast two-hybrid interaction . | β-Galactosidase assay (± one SD)b . |
|---|---|---|---|
| HS11AA | Positive | Positive | ND |
| GE36AA | Positive | Positive | ND |
| S47A | Positive | Positive | ND |
| EF48AA | Positive | Positive | ND |
| S52A | Positive | Positive | ND |
| S55A | Positive | Positive | 449 ± 139 |
| GLI58AAA | Positive | Negative | 2.7 ± 0.8 |
| SE67AA | Positive | Negative | 1.8 ± 0.9 |
| DI69AA | Positive | Negative | 1.2 ± 0.4 |
| SE79AA | Positive | Positive | 131 ± 41 |
The assay of direct interactions between hZwint-1 and hZW10 site-directed mutagenesis constructs analyzed with the LexA yeast two-hybrid system using Xgal and the β-galactosidase assay, as well as kinetochore localization, through fluorescence microscopy analysis of EGFP fusion constructs.
The point mutations were verified by sequencing.
The hZwint-1 interaction with hZw10 point mutants were assayed as outlined in Table I.