Structure elucidation by diffraction methods – contribution to structural biology
J. Dohnálek
Institute
of Macromolecular Chemistry of the Academy of Science of the Czech Republic,
Heyrovského nám. 2, 16206 Praha 6, Czech Republic
dohnalek@imc.cas.cz
Keywords: protein crystallography, X-ray
diffraction, structural biology
The first
documented X-ray scattering on ordered samples of biological origin was
observed by J. D. Bernal in Cambridge, U.K. in 1934. Further experiments by
Dorothy Crowfoot-Hodgkin and M. F. Perutz in Oxford on crystals of insulin and
haemoglobin in 1937 and in the following years lead to development of a method
which revolutionized our views of biochemical processes and molecular structure
of living systems. The number of structures determined by diffraction methods
from 1960 to these days (the first one of myoglobin by Kendrew) amounts to tens
of thousands, many of them remaining unpublished or subject to proprietary
rights of private companies. The length of the process of solving and
finalizing a structure dramatically decreased from several years to several
weeks or months depending on the project difficulty.
While from
1950s till 1980s methods for structure solving such as molecular replacement
and isomorphous replacement were developed, late 1980s and 1990s saw a fast
development of computational technology and its application in all steps of
macromolecular structure determination. In the recent ten years this field
profited from huge development effort in crystallographic software as well as
methodological advances in protein and nucleic acid crystallization and in data
collection and analysis. Macromolecular crystallography today would be
unthinkable without cryo-cooling of samples, synchrotron sources of high
intensity X-ray radiation and high speed internet services and databases.
Crystallographically
determined structures of biological molecules bring invaluable information
ranging from details of interactions between ligands and enzymes, assemblies of
protein-protein or nucleic acid – protein types to highly complex structures of
viruses. Quality of acquired structural information does not depend on the size
of the system once regularly organized.
The most
recent advances (5 years) target the main bottlenecks of these methods and
indicate future development. High throughput methods for protein production,
purification and crystallization, robotized home source and synchrotron
beam-lines and software equipment for automatic information-with-sample
transfer and for automatic data collection were produced mainly within
structural genomics projects for which the main starting impulse were the
results of successful genomics projects. Systematic studies on radiation damage
of biological samples induced by intensive radiation sources contributed
substantially to better experiment planning and techniques. Free electron laser and single particle
imaging are becoming centres of interest mainly for diffraction studies of
larger systems (organelles or cells) without the need of an ordered crystalline
system.
Acknowledgements.
Introduction
of modern methods is supported by the Ministry of Education of the Czech
Republic (1K05008).