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The Growth and Preliminary Investigation of Protein and Nucleic Acid Crystals for X‐ray Diffraction Analysis

Published on Oct 31, 2006in Methods of biochemical analysis
· DOI :10.1002/9780470110430.ch4
Alexander McPherson4
Estimated H-index: 4
(Penn State Milton S. Hershey Medical Center)
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Abstract
  • References (212)
  • Citations (169)
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References212
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#1Alexander McPherson (Penn State Milton S. Hershey Medical Center)H-Index: 4
#2Roderick Spencer (MIT: Massachusetts Institute of Technology)H-Index: 1
Abstract The plant seed protein canavalin has been crystallized alter trypsinization by using vapor diffusion to effect isoelectric precipitation. The space groups and cell dimensions are R3 with a = 81.3 A , γ = 111 ° and having extremely high R 32 pseudo symmetry, P6 3 with a = 126 A and c = 51 A , C222 1 , with a = 136 A , b = 152 A , and c = 131 A . X-ray data combined with electron microscopy suggests the molecule to be composed of six identical subunits. each of 18,500 daltons, arranged in...
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#1Carolyn Cohen (Brandeis University)H-Index: 47
#2Nancy M. TooneyH-Index: 5
MOST fibrous proteins do not crystallise. An artful way to promote protein crystallisation is to modify the structure by proteolytic attack. For example, papain cleavage of γ-globulin yields the homogeneous Fc fragment which readily crystallises1. Following a similar approach, we have crystallised fibrinogen.
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#1G. Zaloga (SBU: Stony Brook University)H-Index: 1
#2R. Sarma (SBU: Stony Brook University)H-Index: 1
A CONSIDERABLE problem hampering the analysis of protein crystal structures has been the damage caused to the crystals by the X-ray beam. Another limiting factor has been the resolution in the diffraction pattern. To increase the useful life of a protein crystal in the X-ray beam1, single crystal monochromators and low temperatures have been used. We now report that styrene and other vinyl monomers can decrease the radiation damage of the crystal in the X-ray beam by a factor of as much as 10, a...
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X-ray crystallographic studies of hemerythrin from Golfingia gouldii show that the molecules pack in a tetragonal cell with two sets of molecules in an apparently face-centred array seen in projection along the 4-fold axis, but displaced relatively to each other by approximately c10 in the axial direction. Both sets of molecules lie on 4-fold rotation axes, so that the subunits of each octameric molecule are related in pairs by a molecular 4-fold rotation axis. The two subunits of each pair are ...
7 CitationsSource
#1John L. Wang (Rockefeller University)H-Index: 9
#2Joseph W. Becker (Rockefeller University)H-Index: 6
Last. Gerald M. Edelman (Rockefeller University)H-Index: 132
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Abstract A lectin from the fava bean ( Vicia faba ) has been purified and crystallized in a form suitable for high-resolution crystallographic structure analysis. This protein binds glucose- and mannose-like saccharides, and it is mitogenic for lymphocytes. The fava lectin crystallizes in the orthorhombic space group. P 2 1 2 1 2 1 with unit cell dimensions a = 90.0, b = 89.3, and c = 67.4 A . The mass of protein in the asymmetric unit is 53,000 daltons, corresponding to the molecular weight of ...
38 CitationsSource
#1Jon D. RobertusH-Index: 43
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The structure of a tRNA has been determined by isomorphous replacement. Some of the interactions which maintain the tertiary structure are of a novel type. Our model differs significantly from one which has recently been proposed.
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#1Sung-Hou Kim (Duke University)H-Index: 84
#2F. L. Suddath (MIT: Massachusetts Institute of Technology)H-Index: 12
Last. Alexander Rich (MIT: Massachusetts Institute of Technology)H-Index: 101
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The 3-angstrom electron density map of crystalline yeast phenylalanine transfer RNA has provided us with a complete three-dimensional model which defines the positions of all of the nucleotide residues in the moleclule. The overall features of the molecule are virtually the same as those seen at a resolution of 4 angstroms except that many additional details of tertiary structure are now visualized. Ten types of hydrogen bonding are identified which define the specificity of tertiary interaction...
702 CitationsSource
#1Christine Schubert Wright (Princeton University)H-Index: 6
#2C. Keith (Princeton University)H-Index: 1
Last. M.M. Burger (University of Basel)H-Index: 1
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Abstract Wheat germ agglutinin crystallizes in two monoclinic space groups, P 2 1 and C 2, under identical crystallization conditions. Unit cell dimensions are a = 73.8 A , b = 51.2 A , c = 90.8 A , γ = 90 ° for P2 1 ; a = 51.31 A , b = 73.35 A , c = 91.45 A , β = 97.75 ° for C 2, both with eight subunit molecules in the unit cell. The C 2 crystals were chosen as suitable for investigating the three-dimensional structure to high resolution, because of their smaller asymmetric unit (containing th...
17 CitationsSource
Abstract A Bence-Jones protein, Rhe, isolated from the urine of a myeloma patient, has been crystallized. Immunological tests indicate that Rhe contains the constant region of a lambda-type light chain. Gel filtration and sodium dodecyl sulfate gel electrophoresis show that Rhe consists of two subunits, each about half the size of the light chain. Preliminary X-ray crystallographic results and density measurements show that Rhe possesses crystallographic 2-fold symmetry, indicating that the two ...
13 CitationsSource
Abstract Single crystals of the yeast enzyme inorganic pyrophosphatase that are suitable for x-ray analysis have been grown. The space group of the crystals is P21. Their unit cell dimensions are: a = 70.4, b = 95.4, c = 52.2 A, and β = 99.8°. There is 1 dimeric enzyme molecule with a molecular weight of approximately 68,000 in the asymmetric unit of the unit cell. The crystal is approximately 57% protein by weight.
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Abstract Salt/polyethylene glycol (PEG) mixtures are employed as precipitants for biological macromolecules. The dependence of precipitation curves (PCs) on salt species was investigated for integral membrane protein/detergent particles. By relating this dependence to properties of ions dissociated from added salts, the following roles and effects of various ions were clarified. In the presence of ions whose interaction with water is stronger than water–water interaction, the coordination of sol...
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#1Claudia Fasolato (University of Perugia)H-Index: 2
An interesting approach to the study of matter is represented by spectroscopy, that infers information about materials and biosystems by driving their interaction with controlled probes, as beams of light, electrons and neutrons. In this chapter, the basic physics of Raman and Surface Enhanced Raman Scattering (SERS) will be discussed. Furthermore, we will present a review of the scientific literature concerning the themes that are relevant for the Thesis, and in particular the fabrication of na...
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Abstract The crystallization of FeS cluster-containing proteins has been challenging due to their oxygen sensitivity, and yet these enzymes are involved in many critical catalytic reactions. The last few years have seen a wealth of innovative experiments designed to elucidate not just structural but mechanistic insights into FeS cluster enzymes. Here, we focus on the crystallization of hydrogenases, which catalyze the reversible reduction of protons to hydrogen, and nitrogenases, which reduce di...
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The crystallization of protein samples remains the most significant challenge in structure determination by X-ray crystallography. Here, the effectiveness of transmission electron microscopy (TEM) analysis to aid in the crystallization of biological macromolecules is demonstrated. It was found that the presence of well ordered lattices with higher order Bragg spots, revealed by Fourier analysis of TEM images, is a good predictor of diffraction-quality crystals. Moreover, the use of TEM allowed (...
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#1Takayuki Odahara (AIST: National Institute of Advanced Industrial Science and Technology)H-Index: 6
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Mixtures of neutral salts and polyethylene glycol are used for various purposes in biological studies. Although the effects of each component of the mixtures are theoretically well investigated, comprehension of their integrated effects remains insufficient. In this work, their roles and effects as a precipitant were clarified by studying dependence of precipitation curves on salt concentration for integral membrane protein/detergent particles of different physicochemical properties. The depende...
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While crystallization historically predates crystallography, it is a critical step for the crystallographic process. The rich history of crystallization and how that history influences current practices is described. The tremendous impact of crystallization screens on the field is discussed.
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The current practice for identifying crystal hits for X-ray crystallography relies on optical microscopy techniques that are limited to detecting crystals no smaller than 5 μm. Because of these limitations, nanometer-sized protein crystals cannot be distinguished from common amorphous precipitates, and therefore go unnoticed during screening. These crystals would be ideal candidates for further optimization or for femtosecond X-ray protein nanocrystallography. The latter technique offers the pos...
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