by Peter Nollert
December 4, 2009 13:00
Many crystallizers love to brag about the 'beast they tamed'. They tell stories describing all the different tools that were used and how in the end a pinch of luck was required to get a recalcitrant protein to crystallize and determine its structure.
How is it possible to judge that one protein is more difficult to deal with than another? Could it be that the standard 'round' approach taken didn't fit the square 'peg'? Could it be that you just followed a pretty but dead-end crystal form while ignoring an early ugly but productive one? Maybe you're not applying the tools properly? Do the fish you're trying to catch swim right through the large-size mesh? You see where I'm going with this: Struggling with a protein crystallization project may have many different reasons, you may be a poor crystallizer or the protein may have a very narrow crystallization slot, that's just very difficult to hit. I guess this could be at the heart of the 'crystallization is more of an art than a science' comments I don't think are useful.
Some protein crystallization projects are just a 'slam dunk'
Regardless, it's a fact that some (not many) proteins express well, can be purified by standard two step chromatography to >95% purity, concentrate well and crystallize in 5% of all formulations of Wizard I, II, III & IV screens and in Cryo I - and diffract to 2A on a home source without any further cryoprotection. Some scientists call these proteins 'well behaved' (when you're a PhD student and your project involves determining a protein structure, my advise is to get several irons in the fire to identify such 'well behaved' protein). Nevertheless, the more proteins are put into structure determination pipelines and the more X-ray structures are determined, the higher the likelihood that you're confronted with targets that are not 'low hanging fruit' and that require a more sophisticated approach than that of a standard structural genomics pipeline. This is the challenging end of the spectrum and when you're done with limited proteolysis, engineering fragments and surface mutants and get a double mutant protein to form well-diffracting crystals you're welcome to exercise your bragging rights by publishing a paper titled "The taming of...".
This is exactly what Baranova et al. have done in this month's edition of Acta Cryst:
E. V. Baranova, S. Beelen, N. B. Gusev and S. V. Strelkov
The taming of small heat-shock proteins: crystallization of the [alpha]-crystallin domain from human Hsp27
Acta Cryst. (2009). F65, 1277-1281
You guys are awesome!
Peter