BIOC 511 Exam 2 Protein Structure Methods

HP Chromatography

Primary structure

pro: gives amino acid content

con: does not give information about sequence

Edman Degradation

Primary structure

pro: provides sequential information

con: only first ~30 residues, N-terminal only

Mass spectrometry

Primary structure

pro: can be used w/amino acid sequencing to build protein

con: complex, requires fragmentation, hard to distinguish repeating subunits

CD spectroscopy

Secondary structure

pro: can estimate secondary structure content

con: low image resolution

FTIR

Secondary structure

pro: can be used to identify secondary structure and picks up on CO groups

con:

Solution (Liquid-State) NMR

tertiary structure

pros: structure of soluble proteins, protons easily detected, physiologic temperature, protein dynamics

con: low signal, labeling often required, size limitation

Solid-state NMR

tertiary structure

pro: structure of insoluble proteins, every temperature possible, protein dynamics

con: low signal, labeling often required, magic angle spinning

EPR

tertiary structure

pro: soluble and insoluble proteins, every temperature possible, protein dynamics

con: labeling always required, one sample per site of interestt

Crystallography

tertiary structure

pro: structure of soluble proteins, very little sample required, very high resolution possible, fast once crystal available

con: needs high quality crystal, usually low temps, no protein dynamics, protons difficult to detect

Cryo Em

tertiary structure

pro: soluble and insoluble proteins, very little sample required, no labeling required

con: needs low temps, no protein dynamics, protons difficult to detect

Homology Modelling (alpha fold)

tertiary structure

pro: very inexpensive and quick, high accuracy for globular proteins

con: no information about intrinsic disorder, no information about dynamics, no information about structural plasticity