Table 1. DeNeddylating enzymes in Plasmodium falciparum.
| Protein name | Gene ID | HsNedd8-VS | FLAG-PfNedd8-Prg_1 | FLAG-PfNedd8-Prg_2 | Activity | |||
|---|---|---|---|---|---|---|---|---|
| Spectrum count | Coverage % | Spectrum count | Coverage % | Spectrum count | Coverage % | |||
| PfUCHL3 |
Pf3D7_1460400 |
1 | 8.6 | 5 | 20.3 | 0 | 0 | Ub HsNedd8 PfNedd8 |
| PfUCH37 | Pf3D7_1117100 | 36 | 34.8 | 15 | 17.4 | 5 | 9.7 | Ub HsNedd8 PfNedd8 |
| Ubiquitin carboxyl -terminal hydrolase 13, putative |
Pf3D7_0413900 | 0 | 0 | 0 | 0 | 4 | 5.5 | Ub |
| Ubiquitin carboxyl- terminal hydrolase, putative |
Pf3D7_0726500 | 0 | 0 | 1 | 0.8 | 4 | 1.9 | Unknown (did not express) |
P. falciparum cell lysate was functionally screened for deNeddylases using FLAG-HsNedd8-VS or FLAG-PfNedd8-Prg, the latter was performed twice (FLAG-PfNedd8-Prg_1 and FLAG-PfNedd8-Prg_2). Reactions were followed by anti-FLAG immunoprecipitation and proteomic analysis of bound enzymes. Enzymes were individually expressed in E.coli, purified, and their specificity was validated by reacting with Ub-AMC, HsNedd8-AMC and PfNedd8-AMC.