Midterm: Lecture 8 Protein crystallography Flashcards
What does the diagram on slide 1 show?
A single large crystal of satellite tobacco mosaic virus
What does the second diagram show on slide 1?
TMV coat protein disk
Define protein:
A polymeric macromolecule joined together by peptide bonds
The virus can be crystallized like what?
Salt
What is TMV?
Mosaic pattern on leaf
Several methods are currently used to determine the structure of a protein, including: (3)
- X-ray crystallography
- NMR spectroscopy
- Electron microscopy
What does each method have?
Advantages and disadvantages
In each of these methods, what does the scientist do?
The scientist uses many pieces of information to create the final atomic model
Primarily, the scientist has what?
For X-ray crystallography:
For NMR spectroscopy:
In electron microscopy:
Some kind of experimental data about the structure of the molecule
this is the X-ray diffraction pattern
it is information on the Local Conformation and Distance Between Atoms that are close to one another
It is an image of the overall shape of the molecule
What is NMR spectroscopy more like?
Magnetic resonance imaging
For X-ray crystallography, this is the X-ray diffraction pattern
What does this mean?
Gives idea of how atoms are organized
X-ray crystallography:
For this method,
The protein is purified and and crystallized, then subjected to an intense beam of X-rays
The proteins in the crystal…
Diffract the X-ray beam into one or another characteristic pattern of spots, which are then analyzed (with some tricky methods to determine the phase of the X-ray wave in each spot) to determine the distribution of electrons in the protein
The resulting map of the electron density…
Is then interpreted to determine the location of each atom
What is diffraction?
Spreading of waves as they pass through or around an obstacle (ex: how light diffracts)
What happens in protein crystallography (from what I wrote)?
Purify protein, then shine beam (X-ray), and looking at diffracted pattern, can determine how atoms are organized (structure)
What does Methods for Determining Atomic Structures show? (From what I wrote)
Looking at diffraction patterns, can tell location of atoms
What can X-ray crystallography provide?
Very detailed atomic information, showing every atom in a protein or nucleic acid along with atomic details of ligands, inhibitors, ions, and other molecules that are incorporated into the crystal
However, the process of crystallization…
Is difficult and Can impose limitations on the types of proteins that may be studied by this method
What is X-ray crystallography an excellent method for?
Determining the structures of rigid proteins that form nice, ordered crystals
What are for more difficult to study?
Why?
Flexible proteins
Because crystallography relies on having many, many molecules aligned in exactly the same orientation, like a repeated pattern in wallpaper
Flexible portions of protein…
Will often be invisible in crystallographic electron density maps, since their electron density will be smeared over a large space
Q: What’s a molecular function of protein?
What does this mean?
Enzyme- changes protein confirmation
Protein confirmation: Neurotransmission
Shape: Protein shape is determined by…
Types
The amino acid sequence and how they are … binding
Primary, secondary, tertiary, quaternary
