Hi all,
I am not a crystal structural biologist. I have few questions that could be rather technical. I am aware that there are some proteins that are difficult to study using x-ray crystalllography (e.g small heat shock proteins). What about these proteins that make them hard to study using x-ray crystallography? What are some other general features that makes a protein hard to study using x-ray crystallography? Also if there are some brief reasons at the level of fundamental principles, please feel free to let me know.
Thanks
I may add the following: some proteins, especially membrane proteins, are exceedingly difficult to crystallize, but some researchers have solved this problem by genetically re-engineering the protein. This would involve deleting regions that are known to be disordered, etc. Thus when interpreting 3D crystal structures of re-engineered proteins, one should be very careful to consider the artificial changes that have been made to them.
Hi,
I'm not an expert on X-ray crystallography, but I can say that some of the features which make proteins hard to form good crystals (good enough to study them by X-ray diffraction) include the presence of very flexible regions, or the presence of carbohydrate moieties.
Large, unstable, unsoluble proteins or complexes are hard to prepare and will impede extensive sparse-screening of crystallization conditions.
Structural heterogeneity causes difficulties in crystallization. Dynamic parts of a protein or subunits exchange in a complex, will prevent the formation of regular crystals.
Membrane Proteins.
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2580798/
http://www.cell.com/trends/biochemical-sciences/fulltext/S0968-0004(07)00084-9
https://www.quora.com/Why-is-it-so-difficult-to-find-the-structure-of-transmembrane-proteins
Membrane proteins, which have predominantly hydrophobic surfaces, are hard to crystallize because they tend to aggregate in aqueous solutions. Proteins which have post-translational modifications are also hard to crystallize because the PTMs are usually not uniform among the protein molecules. Some proteins have inherently disordered regions (IDRs) and those are also hard to crystallize.
For some reason proteins with a isoelectric point (pI) around 10 are harder than the ones with the more common pI around 6. Either covalently modify lysine primary amines or try to find a partner for a protein complex.
Proteins that form heterocomplexes can be hard to crystallise on their own. Though sometimes the complexes are harder to crystallise.
Most intrinsically disordered proteins (IDP) are impossible to crystallise.
Although, there might be general notions which proteins are more likely to form crystals compared to other proteins, in my opinion is still hard to predict whether a protein will crystallize or not. Bottom line is that crystallography is an empirical science and one needs to give it a shot before they can conclude that the protein of interest is difficult to crystallize.
Transmembrane and Membrane proteins basically are the most difficult proteins to crystallizeCrystallizing the protein is quite a bit more difficult too. Membrane proteins in detergent micelles which may or may not be charged themselves look like oily blobs with hopefully a domain or two of folded proteins sticking out of them. Compared to a soluble protein with a nice ordered surface in every direction instead of a detergent micelle which might undergo a phase change at high concentration or by the addition of a salt or change in pH, membrane proteins take a task that might take thousands of trials and adds new dimensions to worry about.
Proteins with high surface hydrophobicity and high surface charge are not good candidates for forming crystals in solutions.
Lots of great advice is above... the only thing I see missing from this list is that proteins which have a high degree of flexibility in their structure can be difficult to crystalize because they are inherently disordered and will not form a crystal lattice well. However, if treatments with ligand or inhibitor or partner proteins will reduce the flexibility they can become better crystalizers.
I may add the following: some proteins, especially membrane proteins, are exceedingly difficult to crystallize, but some researchers have solved this problem by genetically re-engineering the protein. This would involve deleting regions that are known to be disordered, etc. Thus when interpreting 3D crystal structures of re-engineered proteins, one should be very careful to consider the artificial changes that have been made to them.
- Badges
- Science topic
- Similar topics
- Biological Science
- Molecular Biology
- Biomolecules
- Proteins