Table 1.
Single-Molecule Protein Complex Studies
| Labeling-Based Selective Observation | ||||||
|---|---|---|---|---|---|---|
|
| ||||||
| Interactions | Systems | Cell extracts | Labeled objects | Labeling by | Substrates | Refs |
| Spliceosome | Yeast | - | - | RNA | 26, 27 | |
| Protein- | Spliceosome | Yeast | NTC, snRNAs (U1, U2, U5) | DHFR-, SNAP-mediated | RNA | 14 |
| Nucleic Acid | Replisome | Xenopus egg § | DNA duplication products | Fluorescent DIG antibody | lambda DNA | 15, 42 |
| Replisome | Xenopus egg § | DNA clamp (PCNA) | PCNA antibody ++ | lambda DNA | 20 | |
| Replisome | Xenopus egg § | Fen1 | mKikGR | lambda DNA | 45 | |
| DNA polymerase * | E. coli | Polymerases (PolBI, DinB) | FGE-mediated | DNA | 35 | |
| Immobilization-Based Selective Observation | ||||||
|---|---|---|---|---|---|---|
|
| ||||||
| Interactions | Systems | Cell extracts | Immobilized objects | Immobilization by | Substrates/Partners | Refs |
| Protein- | Poly(U) polymerase complex | Human §§ | FLAG-mCherry-tagged TUT4 | RFP antibody + | RNA | 16 |
| Nucleic Acid | Helicase * | E. coli | His-tagged PcrA | His antibody + | DNA | 17 |
|
| ||||||
| Replisome | Human | T7-tagged ORCA | T7 antibody + | YFP-Orc1, Cdt1, Geminin | 49 | |
| Protein kinase | Human | FLAG-mCherry-tagged PKA | FLAG antibody + | YFP-PKA | 17 | |
| Protein kinase holoenzyme | Human | PKA | PKA antibody ++ | AKAP | 17 | |
| Protein- | Protein kinase holoenzyme | Human | FLAG-tagged SKIP | FLAG antibody + | YFP-PKA | 48 |
| Protein | Peptide-loading complex | Human | TAP1 | TAP1 antibody+ | TAP2, YFP-Tapasin | 50 |
| Membrane protein complex | Human | YFP-tagged β2AR | YFP antibody + | - | 17 | |
| Mitochondrial protein | Human | YFP-tagged MAVS | YFP antibody + | - | 17 | |
| mTOR signaling complex | Human | FLAG-tagged mTOR | FLAG antibody + | HA-Raptor +++ | 17 | |
*
The system of interest was not a protein complex.
§
fractionated
§§
immunoprecipitated
+
biotinylated
++
immobilized via biotinylated secondary antibody
+++
probed with fluorescent secondary antibody
AKAP: A-kinase anchoring protein
β2AR: β2-adrenergic receptor
DHFR: dihydrofolate reductase
DIG: Digoxigenin
Fen1: flap endonuclease 1
FGE: formylglycine-generating enzyme
NTC: Prp19-complex
PCNA: proliferating cell nuclear antigen
RFP: red fluorescent protein
SKIP: sphingosine kinase interacting protein
SNAP: a variant of O6-alkylguanine-DNA-alkyltransferase
snRNA: small nuclear RNA
TAP: transporter associated with antigen processing
TUT4: terminal uridylyl transferase 4
YFP: yellow fluorescent protein