PROTEIN IDENTIFICATION METHODS

X ray crystallography

most widely used method to solve protein structure BUT requires the molecules be rigid

Diffraction pattern

this was identifed in the 1930’s but unable to be made sense of until 1950, and its a ray of dots to identify protein structure

Determine electron density

the second step in xray crystallography

Myoglobin and hemoglobin

the first protein crystal structures

John Kendrew and Max Perutz

who solved the structure of myoglobin and hemoglobin

NMR spectroscopy

uses chemical shifts and Nuclear overhauser effects to identify atom orientation and solve protein structures

Nuclear overhauser effects

NOEs

H1 NOSEY spectrum

use a table with x’s to identify 2 residues closeness together, which helps build potential structures (mulitple orientations may be correct)

Xray crystallography

protein forms crystalline environment, atoms placed by fitting directly to electron density and the positions are fairly ambigous but can asses both small and large strucutres

NMR spectroscopy

protein in an aqueous environment and typically isotopically labels, id distance between atoms, report multiple conformations, CAN’T do too big of protein bc their signals will bleed together

CryoEM

can use dynamic structures without and large molecules as different conformations can be observed SIMULTANEOUSLY

Computer prediction

take a sequence and predict a protein sequence, David Baker’s group is good at this

AlphaFold2

uses deep learning AI and information gained by humans in the PDB to solve structures of proteins based on already identified proteins,,,,, this can predict quaternary structures