Obtaining 3D Structures of Biomolecules

A collection of question and answer flashcards based on the lecture notes about obtaining 3D structures of biomolecules, particularly focusing on X-ray crystallography, NMR, and electron microscopy.

What are the three main techniques used to experimentally determine protein structures?

X-ray crystallography, Nuclear Magnetic Resonance (NMR) spectroscopy, and cryo-electron microscopy.

What is the primary limitation of X-ray crystallography?

It requires obtaining high-quality single crystals, which is often a difficult and time-consuming process.

What does the term 'supersaturation' refer to in the context of protein crystallization?

It refers to the state achieved when the concentration of purified protein is maximized, allowing for protein aggregation and crystallization.

What is the purpose of the Fourier Transform in X-ray crystallography?

It converts X-ray diffraction data into electron density maps.

What is the main advantage of NMR spectroscopy over X-ray crystallography?

NMR does not require the sample to be crystallized, allowing for the study of proteins in solution.

What is the role of precipitating agents in protein crystallization?

Precipitating agents drive proteins to neutralize surface charges and can lead to either amorphous precipitates or crystals.

What does 'R-factor' measure in the context of macromolecular structures?

It measures the agreement between observed and calculated structure factors, indicating the quality of the structural model.

What is the purpose of the phase recovery techniques in X-ray crystallography?

To recover the lost phase information essential for constructing accurate electron density maps.

Why is electron microscopy (EM) advantageous for studying large biological complexes?

EM can provide structural data for large complexes that may be challenging or impossible to analyze with other techniques.

What challenge does NMR face when analyzing larger proteins?

NMR is typically limited to proteins smaller than 300 residues due to spectral overlap and signal complexity.