Table 1.

Summary of techniques described in this review and their contributions in ISB.

	Technique	Description in the context of ISB	References
Structural characterization of proteins
Common technique	Macromolecular crystallography	•Captures atomic resolution detail of stable protein conformations •Data can be fit into SAXS/molecular docking/low-resolution cryo-EM data	[15]
	NMR	•Captures atomic detail of small, flexible proteins •Data can be fit into SAXS/molecular docking/low-resolution cryo-EM data	[22], [24], [25]
	SAXS/SANS	Provides overall protein complex shape that can be fit with atomic structures	[40], [42], [43], [44]
Recent advancement	Cryo-EM SPA	•Captures high-resolution stable and lower-resolution flexible protein conformations that can be fit into SAXS/molecular docking •Data can provide overall shape to be fit with atomic structures	[45], [52]
	Computational modeling	Detailed atomic subunit predictions which can be fit into SAXS/molecular docking or used in MX analysis	[61], [66], [68]
Identification and characterization of protein–protein interactions
Common technique	Co-IP	Isolates strong protein interactions using affinity pulldowns that can be characterized by MS	[84]
	FRET	Determines domain positioning or how two proteins interact based on proximity of two fluorophores	[101]
Recent advancement	XL-MS	Captures strong and weak interacting partners and identifies which residues are in proximity to each other	[85], [115], [116], [117]
	Molecular docking	Uses structural data and surface predictions to determine how protein complexes interact	[87], [88], [89], [90]
	Proximity labeling	Identifies proteins that come within 10 nm of the protein of interest	[86]
Contextualization of protein–protein interactions
Future of ISB	Whole-cell cryo-ET	•Determines nanometer-scale resolution of how proteins are spatially separated •Provides a snapshot of transient interactions at the time of freezing	[103], [104]
	Single-cell cryo-EM	•Identifies transient and stable protein complexes in a cell •Atomic resolution can be reached if there are multiple copies of the complex	[111], [112]
	XL-MS and cryo-EM SPA	Traps transient and stable protein complexes with crosslinkers which can be characterized with cryo-EM SPA	[102], [114]