How the structure of lysozyme was actually determined

Blake, C. C. F.; Fenn, R. H.; Johnson, L.N.; Koenig, D. F.; Mair, G L R; North, A. C. T.; Oldham, J. W. H.; Phillips, David; Poljak, Roberto J.; Sarma, V. R.; Vernon, C. A.

doi:10.1107/97809553602060000899

Cited by 8 publications

(12 citation statements)

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“…The proteins selected for testing the SeU as a phasing vehicle were: HEW lysozyme16, thaumatin17, bovine trypsin18, cyan fluorescent protein (CFP)19 and histidinol phosphate phosphatase (HPP)20. In addition, a B-DNA Dickerson-Drew dodecamer (DDD)21 was used.…”

Section: Methodsmentioning

confidence: 99%

“…The “classic” approach, used since the early days of protein crystallography, involves derivatization of native crystals by prolonged soaking in solutions or co-crystallization with various reagents containing heavy metals3, such as Hg, Pt, Au etc. Variations of this approach involve, for example, the use of the heavy-metal clusters4, especially suitable for structures of very large macromolecular complexes, the gaseous xenon or krypton pressurized into native crystals5, or the short soaking in salts of halides6 (Br or I).…”

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confidence: 99%

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Selenourea: a convenient phasing vehicle for macromolecular X-ray crystal structures

Luo

2016

Sci Rep

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Majority of novel X-ray crystal structures of proteins are currently solved using the anomalous diffraction signal provided by selenium after incorporation of selenomethionine instead of natural methionine by genetic engineering methods. However, selenium can be inserted into protein crystals in the form of selenourea (SeC(NH2)2), by adding the crystalline powder of selenourea into mother liquor or cryo-solution with native crystals, in analogy to the classic procedure of heavy-atom derivatization. Selenourea is able to bind to reactive groups at the surface of macromolecules primarily through hydrogen bonds, where the selenium atom may serve as acceptor and amide groups as donors. Selenourea has different chemical properties than heavy-atom reagents and halide ions and provides a convenient way of phasing crystal structures of macromolecules.

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Section: Methodsmentioning

confidence: 99%

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confidence: 99%

Selenourea: a convenient phasing vehicle for macromolecular X-ray crystal structures

Luo

2016

Sci Rep

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“…This method was used for solving the first X-ray structures of hemoglobin [2, 3], myoglobin [4] and in other pioneering works. A very illuminating account of the process of crystal structure determination of lysozyme in the early 1960s was presented by Blake et al [5]. In the early days, when diffraction data were recorded on photographic films, their accuracy was only sufficient to utilize the isomorphous signal of heavy-atom derivatives, with differences between reflection intensities of the native and derivative crystals amounting sometimes to as much as 15–25%.…”

Section: Introductionmentioning

confidence: 99%

Many Ways to Derivatize Macromolecules and Their Crystals for Phasing

Dauter

2017

Methods in Molecular Biology

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Due to the availability of many macromolecular models in the Protein Data Bank, the majority of crystal structures are currently solved by molecular replacement. However, truly novel structures can only be solved by one of the versions of the special-atom method. The special atoms such as sulfur, phosphorus or metals could be naturally present in the macromolecules, or could be intentionally introduced in a derivatization process. The isomorphous and/or anomalous scattering of X-rays by these special atoms is then utilized for phasing. There are many ways to obtain potentially useful derivatives, ranging from the introduction of special atoms to proteins or nucleic acids by genetic engineering or by chemical synthesis, to soaking native crystals in solutions of appropriate compounds with heavy and/or anomalously scattering atoms. No approach guarantees the ultimate success and derivatization remains largely a trial-and-error process. In practice, however, there is a very good chance that one of a wide variety of the available procedures will lead to successful structure solution.

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“…All proteins used for structural analysis were initially obtained directly from biological sources, such as hemoglobin from horse blood [4], myoglobin from sperm whale muscles [5], or lysozyme from chicken eggs [21]. These proteins were subsequently crystallized by precipitation from bulk solutions.…”

Section: Introductionmentioning

confidence: 99%

“…26.1.3.2 in ref. [21]). Electron density maps were plotted by hand or, later, with computer-controlled plotters on sheets of transparencies or plexiglass and stacked in the so-called Richards boxes [102] for interpretation by eye of polymer chains in three dimensions.…”

Section: Introductionmentioning

confidence: 99%

Progress in protein crystallography

Dauter

Wlodawer²

2016

PPL

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Macromolecular crystallography evolved enormously from the pioneering days, when structures were solved by “wizards” performing all complicated procedures almost by hand. In the current situation crystal structures of large systems can be often solved very effectively by various powerful automatic programs in days or hours, or even minutes. Such progress is to a large extent coupled to the advances in many other fields, such as genetic engineering, computer technology, availability of synchrotron beam lines and many other techniques, creating the highly interdisciplinary science of macromolecular crystallography. Due to this unprecedented success crystallography is often treated as one of the analytical methods and practiced by researchers interested in structures of macromolecules, but not highly competent in the procedures involved in the process of structure determination. One should therefore take into account that the contemporary, highly automatic systems can produce results almost without human intervention, but the resulting structures must be carefully checked and validated before their release into the public domain.

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How the structure of lysozyme was actually determined

Cited by 8 publications

References 0 publications

Selenourea: a convenient phasing vehicle for macromolecular X-ray crystal structures

Selenourea: a convenient phasing vehicle for macromolecular X-ray crystal structures

Many Ways to Derivatize Macromolecules and Their Crystals for Phasing

Progress in protein crystallography

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