Heavy-atom derivatization

Garman, Elspeth F.; Murray, James W.

doi:10.1107/s0907444903012794

Cited by 58 publications

(47 citation statements)

References 33 publications

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“…For this to be successful, the anomalous scatterer needs to provide sufficient phasing power. Predicting the strength of an anomalous signal can be useful in deciding what levels of incorporation of heavy atom will be minimally required for a successful phasing experiment (Garman & Murray, 2003; http://skuld.bmsc.washington.edu/ scatter/AS_signal.html). In general, the heavier the atom the larger the signal and the easier it will be to measure.…”

Section: Occupancymentioning

confidence: 99%

Selenium incorporation using recombinant techniques

Walden

2010

Acta Crystallogr D Biol Crystallogr

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Using selenomethionine to phase macromolecular structures is common practice in structure determination, along with the use of selenocysteine. Selenium is consequently the most commonly used heavy atom for MAD. In addition to the well established recombinant techniques for the incorporation of selenium in prokaryal expression systems, there have been recent advances in selenium labelling in eukaryal expression, which will be discussed. Tips and things to consider for the purification and crystallization of seleno-labelled proteins are also included.

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Section: Occupancymentioning

confidence: 99%

Selenium incorporation using recombinant techniques

Walden

2010

Acta Crystallogr D Biol Crystallogr

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“…On the other hand, besides the traditional derivatization approaches for protein phasing by heavy-metal cation soaking and co-crystallization, the protein selenomethionine derivatization strategy has been established [36 -39]. The powerful magic seven derivatives and a useful heavy-atom databank have also been established for protein crystallography [40]. The common phasing methods for macromolecule crystallography are the molecular replacement, multiple and single isomorphous replacement (MIR and SIR), multiple and single isomorphous replacement with anomalous scattering (MIRAS and SIRAS, resp.…”

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confidence: 99%

Selenium Derivatization of Nucleic Acids for X‐Ray Crystal‐Structure and Function Studies

Sheng

Huang

2010

Chemistry & Biodiversity

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It is estimated that over two thirds of all new crystal structures of proteins are determined via the protein selenium derivatization (selenomethionine (Se-Met) strategy). This selenium derivatization strategy via MAD (multi-wavelength anomalous dispersion) phasing has revolutionized protein X-ray crystallography. Through our pioneer research, similarly, Se has also been successfully incorporated into nucleic acids to facilitate the X-ray crystal-structure and function studies of nucleic acids. Currently, Se has been stably introduced into nucleic acids by replacing nucleotide O-atom at the positions 2', 4', 5', and in nucleobases and non-bridging phosphates. The Se derivatization of nucleic acids can be achieved through solid-phase chemical synthesis and enzymatic methods, and the Se-derivatized nucleic acids (SeNA) can be easily purified by HPLC, FPLC, and gel electrophoresis to obtain high purity. It has also been demonstrated that the Se derivatization of nucleic acids facilitates the phase determination via MAD phasing without significant perturbation. A growing number of structures of DNAs, RNAs, and protein-nucleic acid complexes have been determined by the Se derivatization and MAD phasing. Furthermore, it was observed that the Se derivatization can facilitate crystallization, especially when it is introduced to the 2'-position. In addition, this novel derivatization strategy has many advantages over the conventional halogen derivatization, such as more choices of the modification sites via the atom-specific substitution of the nucleotide O-atom, better stability under X-ray radiation, and structure isomorphism. Therefore, our Se-derivatization strategy has great potentials to provide rational solutions for both phase determination and high-quality crystal growth in nucleic-acid crystallography. Moreover, the Se derivatization generates the nucleic acids with many new properties and creates a new paradigm of nucleic acids. This review summarizes the recent developments of the atomic site-specific Se derivatization of nucleic acids for structure determination and function study. Several applications of this Se-derivatization strategy in nucleic acid and protein research are also described in this review.

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“…There are many excellent reviews of the subject of HA derivatization, including Blundell & Johnson (1976), Boggon & Shapiro (2000), Garman & Murray, (2003) and Lu & Sun (2014). Useful websites and databases are listed at the end of x2.…”

Section: Which Has To Usementioning

confidence: 99%

An overview of heavy-atom derivatization of protein crystals

Pike

Garman

Krojer

et al. 2016

Acta Crystallogr D Struct Biol

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Heavy-atom derivatization is one of the oldest techniques for obtaining phase information for protein crystals and, although it is no longer the first choice, it remains a useful technique for obtaining phases for unknown structures and for low-resolution data sets. It is also valuable for confirming the chain trace in lowresolution electron-density maps. This overview provides a summary of the technique and is aimed at first-time users of the method. It includes guidelines on when to use it, which heavy atoms are most likely to work, how to prepare heavy-atom solutions, how to derivatize crystals and how to determine whether a crystal is in fact a derivative.

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Heavy-atom derivatization

Cited by 58 publications

References 33 publications

Selenium incorporation using recombinant techniques

Selenium incorporation using recombinant techniques

Selenium Derivatization of Nucleic Acids for X‐Ray Crystal‐Structure and Function Studies

An overview of heavy-atom derivatization of protein crystals

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