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The weak hydrogen bond was first identified in 1935, but it was only in the early 1990s that it really permeated into the consciousness of chemists and biologists. It only seems natural that this interaction was explored first using spectroscopy, followed by crystallography. In structural supramolecular chemistry, a crystal structure is often not the result of hierarchic interaction preferences but a convolution of a large number of strong and weak interactions, each of which affect the rest intimately. Methods for codification of crystal structures must take this into account if they are to be accurate and useful. Of course, the goal of a subject like crystal engineering is to design systems where the interaction preferences are hierarchic, or in other words where the interaction interference is at a minimum. However, most crystal structures are not so predictable and the challenge posed by weak hydrogen bonding effects to the dogma of crystal engineering remains a real one.

This chapter discusses applications of the Pilot ACE. These applications include linear equations, photogrammetry, traffic simulation, matrix programmes, demonstration programmes, and crystallography. The Pilot ACE was a highly successful computer, earning around £100,000 in its four years of life.

This chapter focuses on Austrian-born molecular biologist Max Perutz (1914–2002). Perutz was one of twenty scientific refugees from continental Europe who went on to win Nobel Prizes. A chemist and molecular biologist, he led the first successful attempt to discover the three-dimensional structure of protein molecules using X-ray crystallography, for which he shared the 1962 Nobel Prize. He was the founding chairman of the Laboratory of Molecular Biology in Cambridge, an institution that continues to thrive and counts thirteen Nobel Prize-winners among those who have spent time in its laboratories. Although Perutz applied to the Society for the Protection of Science and Learning (SPSL) for funding, in the event he did not need their money. His case, however, offers an excellent example of the emotional and practical support SPSL's officers extended to all academics who found themselves in precarious situations in the years following the rise to power of the Nazis in Germany and their subsequent conquest or annexation of neighbouring countries.

In the field of macromolecular crystallography, a renaissance occurred in data collection with the advent of two-dimensional area detectors. These rapidly superseded film cameras and diffractometers fitted with proportional counters as the tools of choice for macromolecular data collection, resulting in a huge increase in collection speeds. This chapter discusses in-house data collection. Data collection systems consist of four major components: a source of X-rays, focusing mirrors (optics), a motor-driven goniostat and crystal viewing assembly, and an area detector for recording diffracted images. The chapter first describes the components that make up this assembly. Before proceeding to discuss X-ray generators, it defines two terms that are widely used in describing the attributes of X-ray generation: flux and brilliance. Flux is the number of photons per second per mrad, and brilliance is the number of photons per second per unit phase space volume (with units photons/s/mm²/millirad²). These are important values when comparing X-ray generators with different filament and focal spots sizes.

This chapter begins with a discussion of the theoretical basis of isomorphous replacement. It then discusses the selection of heavy-atom reagents, heavy atoms and their ligands, preparation of heavy-atom derivatives, determination of heavy-atom positions, and refinement of heavy-atom positions.

This chapter casts the high-throughput automation efforts being developed to meet the needs of Structural Genomics initiatives in the framework of an optimization problem. It gives a general overview on optimization techniques with a bias specifically towards the problem of crystallographic refinement. This picture is extended as the chapter considers model building, program flow control, decision-making, validation, and automation. Finer details of different approaches are provided in a conclusive review of some popular software packages and pipelines.

The number of RNA crystal structures has increased in an exponential manner mainly due to the fact that RNA is increasingly viewed as a predominant part of biological processes such as translation, ribozyme catalysis, and gene regulation, riboswitches, and mRNA-protein interactions, for which the gap in structural knowledge is still deep despite the determination of the crystal structure of the ribosome. Structural studies of the ubiquitous roles of RNA at all levels of cellular processes are starting to be supported by technological developments which enable high-throughput crystallography (HTC). Recently, robotics has entered the field of crystallization. Despite the high cost of these robots, they are highly valuable because they significantly shorten the time to set up experiments as well as multiply the number of possible tests by a 100-fold, just by going to the nanolitre scale in terms of liquid sample handling. They also allow samples to be tested that are too scarce for the usual microlitre-scale techniques. Furthermore, they reduce handling time, which can then be spent on more valuable tasks such as macromolecule purification or structure solving. This chapter presents guidelines to purify and set up RNA oligonucleotides crystallization experiments using a robot. An overview of crystallization robots available on the market will also be given with their advantages and drawbacks.

In the living cell, protein-DNA interactions take place during genetic processes such as chromatin organization, recombination, replication, transcription, and DNA repair. In order to fully understand the mechanism of these processes, study of protein-DNA interactions at atomic level is required. Despite the wealth of knowledge and attempts of several investigators to classify protein-DNA complexes on the basis of structural analysis of well over 200 protein-DNA complexes, the prediction and modelling based on three-dimensional structure or amino acid sequence of a protein and nucleotide sequence of DNA is challenging. X-ray crystallography is the most powerful technique used for structural studies and the development of technology for the production and purification of large quantities of proteins and DNA is vital to its success. Advances in oligonucleotide synthesis have not only provided ease of synthesis of large quantities of pure DNA but also of any required DNA sequence. This has resulted in the crystallization and structure determination of a vast number of DNA oligonucleotides and protein-DNA complexes. Since 1994 there has been a dramatic increase in the number of the protein-DNA complex structures solved annually. The first and most difficult step in the X-ray crystallographic study is the growth of well-diffracting crystals of the protein-DNA complex under investigation. This chapter describes methods employed to purify proteins and DNA for cocrystallization, procedures for crystallization of protein-DNA, and characterization of cocrystals.

This chapter discusses all aspects of the methodology used to solve isometric virus structures with separate case study examples for two demanding examples, the blue tongue virus core (BTV) and bacteriophage PRD1, the first structure of an intact virus with an internal membrane.

This chapter discusses macromolecular crystallography in drug discovery. Currently, all large and most small pharmaceutical and biotechnology companies have invested in macromolecular crystallography, with protein crystallography now an integral tool of drug discovery. During the last twenty years, the term rational drug design has slowly been replaced with the more precise term structure-based drug design (SBDD).

This chapter focuses on the discovery of DNA. The discovery of DNA as the basis of the gene, how it is reproduced, and how it affects protein synthesis was the pinnacle of life sciences in the 20th century. Gene splicing, recombinant DNA, transgenic organisms (or genetically modified plants and animals), and the patenting of genes from microbes to humans all have roots in fundamental discoveries in molecular biology during the 1950s and 1960s. The molecular approach to the gene involved a merger of microbiology and genetics using three techniques from physics and chemistry: (i) radioisotopes were used to help identify DNA as the basis of the gene; (ii) X-ray crystallography was used to reveal the three-dimensional structure of proteins and of DNA; and (iii) chromatography was used to analyze the composition of DNA and proteins.

Fritz Haber hired Polanyi to work in the Fiber Chemistry Group of the Kaiser Wilhelm Institute in Berlin. Polanyi helped develop the rotating-crystal method of X-ray crystallography, made solid contributions to understanding the structure of cellulose, pressed forward with his work on adsorption catalysis and electrostatic dipoles, laid the foundation for transition rate theory in reaction kinetics, and investigated the bond strength of crystals; he was also forced to give up a cherished theory about quantum jumps in reaction kinetics, which taught him an important lesson about how scientists work together to distinguish real discoveries from mistaken surmises. Polanyi married Magda Kemeny on February 21, 1921, in a civil ceremony; their first child, George Michael Polanyi, was born on October 1, 1922.

The historical introduction covers the parallel developments in EM on inorganic and biological samples. Vainshtein, Pinsker and Zvyagin in Moscow solved structures from electron-diffraction patterns in the 1950s. Aaron Klug in 1968 pioneered computer image processing of EM images and realised that the crystallographic structure factor phase information can be read out directly in numbers from the Fourier transform. A few years later the EM optics was powerful enough to allow metals to be seen in oxides. The first determination of atomic coordinates in a crystal from EM images was done in 1984. Compared to X-ray crystallography, electron microscopy has some advantages; crystals a million times smaller (even defects) can be studied and the crystallographic structure factor phases can be read out numerically. Drawbacks are radiation damage (especially for organics) and multiple diffraction.

This introductory chapter begins with an appreciation of the unique position of electron microscopy in biological research as it bridges a wide gap between X-ray crystallography and light microscopy. The scope of this book is defined to cover three-dimensional imaging of molecular assemblies that exist in an in vitro sample in large numbers with identical or near-identical structure. Only with such samples it is possible to collect large numbers of projection images suitable for averaging and three-dimensional reconstruction. The fact that molecules can be imaged as single, isolated particles embedded in ice (“crystallography without crystals”) makes the techniques described in this book uniquely suited to image molecular machines in their various processing states. In sharp contrast, electron tomography, not covered in this book, is concerned with the three-dimensional imaging of “unique” objects that may be an organelle or slice of a cell. The vision of a unified structural analysis of macromolecules is articulated, which would lead to an integration of results from cryo-EM, X-ray crystallography and NMR, and a cross-fertilization among these disciplines. The chapter concludes by making the point that the development of single-particle reconstruction would not have been possible without the vast increase in computer power seen in the past decades.