Table 2.
Comparison between different PL approaches
| Enzyme | Organelles | Strengths | Limitations |
|---|---|---|---|
| APEX2 | Cytosol, nucleus (138), mitochondrial matrix (139), mitochondrial intermembrane space (48), mitochondrial nucleoid (139), ER lumen (140), ERM (141), OMM (142), Golgi (143), autophagosome (144), and cilia (145) | High temporal resolution and high activity in most cellular compartments. Can be used for RNA labeling and electron microscopy | Limited application in vivo because of H2O2 toxicity and low permeability |
| HRP | ER lumen, cell surface (146), and synaptic cleft (54) | Higher activity than APEX2 in the secretory pathway. Used for cell surface proteomics | Limited to secretory pathway and extracellularly; limited use in vivo because of H2O2 toxicity and low permeability |
| BioID | Cytosol, nucleus (147), ERM (148), mitochondrial matrix (149), mitochondrial outer membrane (150), mitochondrial nucleoid (151), cell surface (152), peroxisome membrane (153), Golgi (154), and cilia (155) | Nontoxic labeling conditions and ideal for most cellular compartments | Poor temporal resolution and limited application in vivo because of low catalytic activity |
| BioID2 | Higher activity than BioID. Stable at higher temperatures | ||
| BASU | Cytosol, nucleus (156) | Higher activity than BioID | |
| AirID | Cytosol, nucleus (46) | Higher activity than BioID. Lower potential toxicity in long-term experiments | |
| TurboID | Cytosol, nucleus, ERM, ER lumen, mitochondrial matrix, and cell surface (36) | Highest activity promiscuous biotin ligase | Less user control of labeling window because of high biotin affinity. Toxicity in long-term experiments |
| miniTurboID | High resolution and tight control of user-defined labeling window | Lower catalytic activity and stability compared with TurboID | |
| Split-APEX2 | Cytosol, nucleus, ER–mitochondria contact sites, and cell surface (157) | High reconstituted activity | Limited application in vivo because of reagent toxicity and low reagent permeability; requires supplementation of heme cofactor |
| Split-HRP | ER lumen, cell surface, and neuronal synapse (95) | High reconstituted activity | Limited application in vivo because of reagent toxicity and low reagent permeability; requires supplementation of heme cofactor; limited to secretory pathway and extracellular applications |
| Split-BioID | Cytosol, nucleus, and ER–mitochondria contact sites (57, 58, 59) | Nontoxic labeling conditions | Poor temporal resolution and very low catalytic activity |