X-ray crystallography, cryo-EM, NMR, AlphaFold, protein-structure determination pipelines.
Structural Biology Methods
X-ray crystallography
Determines atomic structure from X-ray diffraction by protein crystal. Phase problem solved by MR, MAD, SAD, direct methods. Resolution:…
Cryo-electron microscopy
Single-particle electron microscopy of vitrified specimens. 'Resolution revolution' (2013): direct electron detectors + 3D reconstruction…
Protein NMR spectroscopy
Protein structure determination from chemical-shift + NOE + RDC restraints. Size limit ~40-50 kDa; unique for dynamics (μs-ms motions,…
AlphaFold
DeepMind 2020/2021 neural-network protein-structure predictor. CASP14 breakthrough; AF2 achieves experimental accuracy on most domains…
Mass spectrometry proteomics
Peptide identification + quantification by MS/MS. Bottom-up (tryptic digest) most common. Workflows: LC-MS/MS → database search (Mascot,…
Biomolecular molecular dynamics
Newtonian integration of atomic motions using force fields (AMBER, CHARMM, GROMOS). Timescales: μs-ms with enhanced sampling (REMD,…
Size-exclusion chromatography + PDB
Protein preparation: express → purify (Ni-NTA, SEC, IEC) → biophysical QC (DLS, CD, SEC-MALS). Structures deposited in Protein Data Bank…
Small-angle X-ray scattering (SAXS)
Solution-scattering technique reporting low-resolution shape + oligomeric state. Pair-distance distribution function P(r), Rg, molecular…
Single-molecule FRET
FRET between donor-acceptor fluorophores on single molecule reveals conformational dynamics. Distance range 2-10 nm. Reveals…
Hydrogen-deuterium exchange MS
Backbone amide H → D exchange rate probes solvent accessibility + dynamics. HDX-MS maps protein-ligand interfaces, allosteric changes,…
Phase retrieval direct methods (Hauptman-Karle)
Hauptman-Karle direct-methods solve crystallographic phase problem from intensity data via Sayre tangent formula and Karle-Hauptman…
Cryo-EM Bayesian 3D reconstruction (RELION)
Scheres 2012 RELION: maximum-a-posteriori reconstruction θ* = argmax p(X|θ)p(θ) on volume θ given particle images X; Wiener-filter…
X-ray (Perutz-Kendrew 1958)
Perutz-Kendrew 1958 hemoglobin + myoglobin crystal-structures (Nobel 1962); modern XFEL serial-femtosecond Chapman 2011.
Solution NMR (Wuthrich 1985)
K Wuthrich 1985 NOESY 2D-NMR + 3D-NMR; Nobel 2002; modern up to ~70 kDa proteins; complementary to X-ray + cryo-EM.
Cryo-EM (Kuhlbrandt 2014)
W Kuhlbrandt 2014 'Resolution Revolution'; direct-detector + Bayesian-RELION; modern 1.2 A Yip 2020 apoferritin record.
AlphaFold 2 (Jumper 2021)
DeepMind 2021 attention-based protein-structure-prediction; ~1A median accuracy; AF3 multi-modal 2024 + RNA + DNA + ligand.
HDX-MS (Engen 1999)
J Engen 1999 hydrogen-deuterium-exchange MS; modern PLM-HDX + millisecond-resolved; conformational dynamics standard.
smFRET (Stryer-Haugland 1967)
Stryer-Haugland 1967 FRET as spectroscopic ruler; modern Ha-Selvin 1996 single-molecule FRET; conformational landscapes.
X-ray crystallography (Perutz-Kendrew 1958)
M Perutz-J Kendrew 1958 (Nobel 1962) Hb / Mb structure; modern modern MX synchrotron + XFEL serial-crystallography + drug-design pipeline.
Cryo-EM revolution (Kühlbrandt 2014)
W Kühlbrandt 2014 'Resolution Revolution' Science; modern modern direct-detector Falcon + Gatan + 2024 1.2Å apo-ferritin.
Bio-NMR (Wüthrich 1985)
K Wüthrich 1985 (Nobel 2002) NOESY-distance-constraints; modern modern TROSY-deuterated + paramagnetic + 1.2-GHz dynamics-fingerprint.
AlphaFold2 (Jumper 2021)
Jumper-Hassabis 2021 AlphaFold2; modern modern AF3 multimer + RoseTTAFold-AllAtom 2024 + ESMFold + protein-design pipeline.
XFEL (Chapman 2011)
H Chapman 2011 first XFEL serial-crystallography photosystem-II; modern modern modern time-resolved + diffraction-before-destruction.
MD simulation (McCammon 1977)
A McCammon 1977 first protein-MD BPTI; modern modern AMBER + GROMACS + Anton + 2024 ML-IF protein-folding-MD generative.