The Importance of Protein Purity and Homogeneity

Timing is also important, and when one is carrying out initial trials it is good to examine the crystallization samples frequently, every 12 to 24 h for the first few days. This way conditions that cause very rapid precipitation or crystal growth can be identified. Once optimal crystal growth conditions have been precisely defined, then that is the time to lay the trials down like fine wine, in a cool, quiet place. 

It is wise to pay attention to what might be considered trivial matters. Be certain that the workplace is clean to minimize dust and microbes in the samples. When making a microdroplet, see that it is as hemispherical as possible and does not spread on the glass or plastic to yield a large surface-to-volume ratio. Microfilter protein samples, work quickly to avoid evaporation do not carry on philosophical conversations while dispensing ingredients. Be alert for unusual events that may later explain anomalous results. Be patient. Examine the volume by McPherson (1999). 

Certainly important to promoting periodic bond formation is that the population of molecules be as homogeneous as possible. For this reason any contamination by unwanted 

There are occasions when even the most intense efforts to crystallize a specific protein fail despite the best efforts at ultra-purification and elimination of microheterogeneity. When this occurs, an alternative is to turn to a different source of the protein. Often only very small variations in amino acid sequence, as found for example between different species of organisms, is enough to produce dramatic differences in the crystallization behavior of a protein. Thus, if the protein from one source proves intractable, consider another. With recombinant proteins, of course, one always has the option of producing a vast range of mutants. Variation of sequence in fact provides a powerful approach to crystallizing proteins when the native molecule fails. There is currently much work underway to define effective mutation strategies for protein crystallization (Derewenda, 2004 Dale et al., 2003). 

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