Solution conformation and crystal structure of methyl 3,6-dideoxy-β-d-ribohexopyranoside,an immunodominant sugar of O-antigens

The crystal structure of methyl 3,6-dideoxy-β-d-ribohexopyranoside monohydrate was determined by direct methods. Crystals are monoclinic, space group P2₁, with cell dimensions $\text{a = 9.089(1), b = 7.668(1), c = 6.956(1)}\text{A}\text{?}\text{, β = 101.12°}$. The molecule adopts the ⁴C₁ chair conformation. The same conformation was also found in both aqueous and chloroform solutions. The pyranose ring is only slightly distorted, and the consequences of this observation on antigen structure are discussed.     

Crystallographic study of plastocyanins

Crystals of plastocyanins from pea and corn leaves have been obtained. Both are suitable for X-ray structure analysis with a resolution up to 1.8 Å. The crystal form of plastocyanin from pea leaves belongs to the space group P2₁2₁2₁ with unit cell dimensions: $\text{a = 49.0}\text{A}\text{?}\text{, b = 53.3}\text{A}\text{?}\text{, c = 82.6}\text{A}\text{?}$. The assumed number of protein molecules per asymmetric unit of the unit cell is two. Crystals of the oxidized (Cu²⁺) and reduced (Cu⁺) forms are isomorphic. No essential differences in spot intensities for the main zone with a resolution of 3 Å were revealed. The crystal form of plastocyanin from corn leaves belongs to the space group P1 with unit cell parameters: $\text{a = 24.8}\text{A}\text{?}\text{, b = 30.0}\text{A}\text{?}\text{, c = 58.5}\text{A}\text{?}$ and α = 96° 10′, β = 87°08′, γ = 78°40′. The assumed number of protein molecules per asymmetric unit is two.     

A preliminary crystallographic study of wheat germ agglutinin

Wheat germ agglutinin crystallizes in two monoclinic space groups, P2₁ and C2, under identical crystallization conditions. Unit cell dimensions are $\text{a = 73.8}\text{A}\text{?}\text{, b = 51.2}\text{A}\text{?}\text{, c = 90.8}\text{A}\text{?}\text{, γ = 90 °}$ for $\text{P2}{}_1\text{; a = 51.31}\text{A}\text{?}\text{, b = 73.35}\text{A}\text{?}\text{, c = 91.45}\text{A}\text{?}\text{, β = 97.75 °}$ for C2, both with eight subunit molecules in the unit cell. The C2 crystals were chosen as suitable for investigating the three-dimensional structure to high resolution, because of their smaller asymmetric unit (containing the dimer), and also because they display better diffraction patterns.     

Preliminary X-ray data for the ribose binding protein from Salmonella typhimurium

Crystals of the periplasmic ribose binding protein of Salmonella typhimurium have been subjected to X-ray analysis. The crystals grow as rectangular parallelopipeds with the symmetry of space group P2₁. Unit cell dimensions are $\text{a = 64·4}\text{A}\text{?}\text{, b = 60·6}\text{A}\text{?}\text{, c = 62·8}\text{A}\text{?}$, and β = 91·25 °. There are two molecules of molecular weight 29,000 per asymmetric unit.     

Highly ordered crystals of the plant seed protein crambin.

Crystals of crambin, a plant seed protein of molecular weight 5000, diffract X-rays strongly to the interplanar spacing limit of 0.88 Å. These diffraction data should allow a definition of atomic structure that is on a par with that typically obtained from crystals of small organic molecules. The crystals are in space group P2₁ and have unit cell dimensions $\text{a = 41.1}\text{A}\text{?}\text{, b = 18.7}\text{A}\text{?}\text{, c = 22·7}\text{A}\text{?}\text{,}\text{and}\text{β = 90.6 °}$. The asymmetric unit contains one protein molecule.     

Crystallographic data for lysozyme from the egg white of the Embden goose

Two crystal forms of lysozyme from the egg white of the embden goose (Anser anser) have been obtained, both of which are suitable for X-ray diffraction analysis. The monoclinic form has space group P2₁ with cell dimensions $\text{a = 38.3}\text{A}\text{?}\text{, b = 65.7}\text{A}\text{?}\text{, c = 45.2}\text{a}\text{?}$, β = 116 ° and the triclinic form (space group P1) has cell dimensions of $\text{a = 39.9}\text{A}\text{?}\text{, b = 42.2}\text{A}\text{?}\text{, c = 57.9}\text{A}\text{?}$ and α = 98.8 °, β = 102.5 °, γ = 90.5 °.     

Structural investigations of mode of action of drugs. I. Molecular structure of mitomycin C.

The crystal and molecular structure of the antitumor antibiotic mitomycin C has been determined by X-ray diffraction. The space group is monoclinic P2₁ with cell dimensions $\text{a}\text{=77.988(3)}$, $\text{b}\text{=20.355(7)}$, $\text{c}\text{=9.679(3)}\text{A}\text{?}$, β=95.99°(1) and Z=4. The structure was solved by direct methods. There are two independent molecules per asymmetric unit. The structure was refined to an R value of 0.049 for 2151 observed reflections measured on diffractometer. The benzoquinone ring is slightly deviated from planarity. The N4 in the indole ring behaves like an amide nitrogen owing to its participation in the conjugated benzoquinoid system. The five membered ring through C1, C2, C3, N4 and C9a adopts an envelope conformation. The molecules are held together in crystal by the hydrogen bonds. All nitrogens except N4 are involved in the hydrogen bonding. Studies with models of drug and DNA indicate two different kinds of mechanism for crosslinking.     

Visualization of drug-nucleic acid interactions at atomic resolution: II. Structure of an ethidium/dinucleoside monophosphate crystalline complex,ethidium:5-iodocytidylyl (3′–5′) guanosine

Ethidium forms a second crystalline complex with the dinucleoside monophosphate 5-iodocytidylyl(3′–5′)guanosine (iodoCpG). These crystals are monoclinic, P2₁, with $\text{a = 14.06}\text{A}\text{?}\text{, b = 32.34}\text{A}\text{?}\text{, c = 16.53}\text{A}\text{?}\text{, β = 117.8 °}$. The structure has been solved to atomic resolution using rigid-body Patterson vector search and Fourier methods, and refined by full matrix least-squares to a residual of 0.16 on 3180 observed reflections. The structure consists of two ethidium molecules, two iodoCpG molecules, 27 water molecules and four methanol molecules, a total of 165 atoms (excluding hydrogens) in the asymmetric unit. Both iodoCpG molecules are hydrogen-bonded together by guanine · cytosine Watson-Crick base-pairing. Adjacent base-pairs within this paired iodoCpG structure and between neighboring iodoCpG molecules in adjoining unit cells are separated by 6.7 Å. This distance reflects the presence of an ethidium molecule intercalated between base-paired iodoCpG molecules and another ethidium molecule stacked above (and below) the dinucleotide. Approximate 2-fold symmetry is used in the interaction; this reflects the pseudo-2-fold symmetry axis of the phenanthridinium ring system in ethidium coinciding with the approximate 2-fold axis relating base-paired iodoCpG molecules. The phenyl and ethyl groups of the intercalated ethidium molecule lie in the narrow groove of the miniature iodoCpG double-helix. The stacked ethidium, however, lies in the opposite direction, its phenyl and ethyl groups neighboring iodine atoms on cytosine residues. Base-pairs within the paired nucleotide units are related by a twist of about 8 °. The magnitude of this angular twist reflects conformational changes in the sugar-phosphate chains accompanying intercalation. These primarily reflect the differences in ribose sugar ring puckering that are observed (i.e. both iodocytidine residues have C3′ endo sugar conformations, while both guanosine residues have C2′ endo sugar conformations), and alterations in the glycosidic torsional angles that describe the base-sugar orientation.The information provided by this structure analysis (along with the accompanying one (ethidium:iodoUpA), described in the previous paper) has led to an understanding of the general nature of intercalative drug binding to DNA. This is described in the third paper of this series.     

Glycerol conformation and molecular packing of membrane lipids: The crystal structure of 2,3-dilauroyl-d-glycerol

The single-crystal structure of 2,3-dilauroyl-d-glycerol has been determined by Patterson rotation and translation methods and refined to R = 0.069. 2,3-dilauroyl-d-glycerol crystallizes in the monoclinic space group P2₁, with unit cell dimensions: $\text{a = 5.46}\text{A}\text{?}\text{, b = 7.59}\text{A}\text{?}\text{, c = 34.2}\text{A}\text{?}\text{and}\text{β = 93.1 °}$, and with two molecules per unit cell. The molecules have their hydrocarbon chains aligned parallel, and are arranged in a bilayer structure. The chain stacking is achieved by a bend in the fatty acid. The hydrocarbon chains pack according to the orthorhombic perpendicular chain packing mode, and are tilted 26.5 ° from the layer normal.The structural features of 2,3-dilauroyl-d-glycerol have been analysed with reference to the corresponding hydrophobic moieties in the crystal structures of different membrane lipids. The glycerol group in 2,3-dilauroyl-d-glycerol is oriented parallel to the layer plane, but changes to an approximately layer-perpendicular orientation when a polar group is attached. The molecular conformation of the glycerol-dicarboxylic ester group, however, is identical in both the absence and presence of a head group, indicating extensive conformational restrictions for this group due to both intrinsic properties and chain stacking. The gathered data provide detailed information on the structural properties of the hydrophobic moiety of membrane lipids.     

Crystallographic data for a penicillin receptor : exocellular DD-carboxypeptidase-transpeptidase from Streptomyces R61.

A pencillin-sensitive enzyme, the exocellular dd-carboxypeptidase-transpeptidase from Streptomyces R61, has been crystallized from polyethylene glycol (M_r = 6000 to 7500) solution at pH 7·6. X-ray examination of the orthorhombic crystals shows the space group is P2₁2₁2₁, with unit cell dimensions $\text{a = 51·1}\text{A}\text{?}\text{, b = 67·4}\text{A}\text{?}\text{,}\text{and}\text{c = 102·9}\text{A}\text{?}$. With four molecules of molecular weight 38,000, the $\text{A}\text{?}{}^3\text{/}\text{dalton}$ ratio for the cell is 2·33. The crystals are stable to irradiation for 75 hours and are suitable for structure analysis to at least 2·4 Å resolution. The radius of gyration of the molecule in solution at pH 6.8 is 20.8 Å.     

Preliminary X-ray diffraction studies on a [4Fe4S] ferredoxin from Bacillus thermoproteolyticus

A [4Fe4S] ferredoxin from Bacillus thermoproteolyticus has been crystallized. The space group is P1 with two molecules in the unit cell, with the dimensions $\text{a = 32.96}\text{A}\text{?}\text{, b = 37.83}\text{A}\text{?}\text{, c = 39.82}\text{A}\text{?}\text{, α = 118.1 °, β = 104.2 °}\text{and}\text{γ = 89.7 °}$. The Bijvoet-difference Patterson map of the native crystal shows up a prominent peak of [4Fe4S] cluster.     

Visualization of drug--nucleic acid interactions at atomic resolution. VI. Structure of two drug--dinucleoside monophosphate crystalline complexes, ellipticine--5-iodocytidylyy (3'-5') guanosine and 3,5,6,8-tetramethyl-N-methyl phenanthrolinium--5-iodocytidylyl (3'-5') guanosine

Ellipticine and 3,5,6,8-tetramethyl-N-methyl phenanthrolinium form complexes with the dinucleoside monophosphate, 5-iodocytidylyl(3′–5′)guanosine. These crystals are isomorphous: ellipticine-iodoCpG² crystals are monoclinic, space group P2₁ with $\text{a = 13.88}\text{A}\text{?}\text{, b = 19.11}\text{A}\text{?}\text{, c = 21.42}\text{A}\text{?}$, β = 105.4; TMP-iodoCpG crystals are monoclinic, space group P2₁, with $\text{a = 13.99}\text{A}\text{?}\text{, b = 19.12}\text{A}\text{?}\text{, c = 21.31}\text{A}\text{?}$, β = 104.9 °. Both structures have been solved to atomic resolution by Patterson and Fourier methods, and refined by full matrix least-squares.The asymmetric unit in the ellipticine-iodoCpG structure contains two ellipticine molecules, two iodoCpG molecules, 20 water molecules and 2 methanol molecules, a total of 144 atoms, whereas, in the tetramethyl-N-methyl phenanthrolinium-iodoCpG complex, the asymmetric unit contains two TMP molecules, two iodoCpG molecules, 17 water molecules and 2 methanol molecules, a total of 141 atoms. In both structures, the two iodoCpG molecules are hydrogenbonded together by guanine-cytosine Watson-Crick base-pairing. Adjacent base-pairs within this paired iodoCpG structure are separated by about 6.7 Å; this separation results from intercalative binding by one ellipticine (or TMP) molecule and stacking by the other ellipticine (or TMP) molecule above or below the base-pairs. Base-pairs within the paired nucleotide units are related by a twist of 10 to 12 °. The magnitude of this angular twist is related to conformational changes in the sugar-phosphate chains that accompany drug intercalation. These changes partly reflect the mixed sugar puckering pattern observed: C3′ endo (3′–5′) C2′ endo (i.e. both iodocytidine residues have C3′ endo conformations, whereas both guanosine residues have C2′ endo conformations), and additional small but systematic changes in torsional angles that involve the phosphodiester linkages and the C4′C5′ bond.The stereochemistry observed in these model drug-nucleic acid intercalative complexes is almost identical to that observed in the ethidium-iodoUpA and -iodoCpG complexes determined previously (Tsai et al., 1975a,b,1977; Jain et al., 1977). This stereochemistry is also very similar to that observed in the 9-aminoacridine-iodoCpG and acridine orange-iodoCpG complexes described in the preceding papers (Sakore et al., 1979 Reddy et al., 1979). We have already proposed this stereochemistry to provide a unified understanding of a large number of intercalative drug-DNA (and RNA) interactions (Sobell et al., 1977a,b), and discuss this aspect of our work further in this paper.     

Polar group interaction and molecular packing of membrane lipids: The crystal structure of lysophosphatidylethanolamine

The conformation and molecular packing of 3-palmitoyl-dl-glycerol-1-phosphoryl-ethanolamine has been determined by a single crystal analysis (R = 0.115); it crystallizes in the monoclinic space group $\text{P2}{}_1\text{a}$ with a unit cell of $\text{a = 7.66}\text{A}\text{?}\text{, b = 9.08}\text{A}\text{?}\text{, c = 37.08}\text{A}\text{?}\text{and}\text{β = 90.2 °}$, with four molecules per unit cell. The molecules exist as configurational and conformational enantiomers and pack in a bilayer arrangement. The phosphorylethanolamine groups have an orientation parallel to the layer surface. The hydrocarbon chains are arranged according to the T∥ chain packing mode and adopt an extreme tilt of 57.5 ° with respect to the layer normal. The free glycerol hydroxyl group forms an intramolecular hydrogen bond with, a phosphate oxygen and thus affects the conformation and orientation of the head group. The phosphorylethanolamine dipoles are oriented parallel to each other in double rows, while they are antiparallel and form a continuous network in dilauroylphosphatidylethanolamine (Elder et al., 1977). The area per molecule in 3-palmitoyl-dl-glycerol-1-phosphorylethanolamine (34.8 Ų) is less than in diacylphosphatidylethanolamine (38.6 Ų), indicating that in the latter the hydrocarbon chains determine the molecular cross-section. The significance of the interaction and space requirement of the phosphorylethanolamine group for the phase behaviour of phosphatidylethanolamine is discussed.     

Crystallization of ribonuclease St

The crystals of ribonuclease St, the extracellular ribonuclease from Streptomyces erythreus, have been obtained from (NH₄)₂SO₄ solution with acetate buffer (pH 4.1). The crystals belong to a monoclinic space group C2 with dimensions $\text{a = 88.4}\text{A}\text{?}\text{, b = 33.0}\text{A}\text{?}\text{, c = 69.0}\text{A}\text{?}\text{, β = 98.4 °}$. There are two protein molecules per asymmetric unit. The crystals diffract beyond 2.0 Å resolution.     

Visualization of drug-nucleic acid interactions at atomic resolution: I. Structure of an ethidium/dinucleoside monophosphate crystalline complex,ethidium:5-Iodouridylyl (3′–5′) adenosine

Ethidium forms a crystalline complex with the dinucleoside monophosphate 5-iodouridylyl(3′–5′)adenosine (iodoUpA). These crystals are monoclinic, space group C2, with unit cell dimensions, $\text{a = 28.45}\text{A}\text{?}\text{, b = 13.54}\text{A}\text{?}\text{, c = 34.13}\text{A}\text{?}\text{, β = 98.6 °}$. The structure has been solved to atomic resolution by Patterson and Fourier methods, and refined by full matrix least-squares to a residual of 0.20 on 2017 observed reflections. The asymmetric unit contains two ethidium molecules, two iodoUpA molecules and 27 water molecules, a total of 155 atoms excluding hydrogens. The two iodoUpA molecules are held together by adenine · uracil Watson-Crick-type base-pairing. Adjacent base-pairs within this paired iodoUpA structure and between neighboring iodoUpA molecules in adjoining unit cells are separated by about 6.7 Å; this separation results from intercalative binding by one ethidium molecule and stacking by the other ethidium molecule above and below the base-pairs. Non-crystallographic 2-fold symmetry is utilized in this model drug-nucleic acid interaction, the intercalated ethidium molecule being oriented such that its phenyl and ethyl groups lie in the narrow groove of the miniature nucleic acid double-helix. Base-pairs within the paired nucleotide units are related by a twist of 8 °. The magnitude of this angular twist is related to conformational changes in the sugar-phosphate chains that accompany drug intercalation. These changes partly reflect the differences in ribose sugar ring puckering that are observed (both iodouridine residues have C3′ endo sugar conformations, whereas both adenosine residues have C2′ endo sugar conformations), and alterations in the glycosidic torsional angles describing the base-sugar orientations. Additional small but systematic changes occur in torsional angles that involve the phosphodiester linkages and the C4′C5′ bond. Solution studies have indicated a marked sequence-specific binding preference in ethidium-dinucleotide interactions, and a probable structural explanation for this is provided by this study.This structure and the accompanying one described in the second paper [ethidium:5-idocytidylyl(3′–5′)guanosine] are examples of model drug-nucleic acid intercalative complexes, and the information provided by their structure analyses has led to a general understanding of intercalative drug binding to DNA. This is described in the third paper of this series.     

A preliminary crystallographic investigation of rabbit muscle creatine kinase

Three crystal forms of rabbit muscle creatine kinase have been grown, one of which seems suited to a high resolution X-ray diffraction study. The first form is of monoclinic space group P2₁ with $\text{a = 54}\text{A}\text{?}\text{, b = 114}\text{A}\text{?}\text{, c = 145}\text{A}\text{?}\text{, β = 91 °}$ and has as the asymmetric unit two molecules of total molecular weight 160, 000. The second form, grown in the presence of mercurials, is of space group A2 with $\text{a = 52}\text{A}\text{?}\text{, b = 165}\text{A}\text{?}\text{, c = 237}\text{A}\text{?}\text{, β = 91 °}$ and also has two molecules in the asymmetric unit. The third crystal form, grown in the presence of a high concentration of cysteine, is of apparent space group P2₁2₁2₁, but evidence indicates that the true space group may be P2₁22₁. The dimensions of the orthorhombic unit cell are $\text{a = 47}\text{A}\text{?}\text{, b = 86}\text{A}\text{?}\text{, c = 125}\text{A}\text{?}$, and the asymmetric unit contains a single protein subunit. Assuming the latter space group, then the creatine kinase molecule possesses a twofold axis relating two identical subunits.     

Disco-ordinate expression of the Escherichia coli gal operon after prophage lambda induction.

EcoRI endonuclease crystallizes in space group C2 with unit cell parameters $\text{a = 209}\text{A}\text{?}\text{, b = 129}\text{A}\text{?}\text{, c = 50}\text{A}\text{?}\text{and}\text{β = 98.4 °}$. Four 29,000 molecular weight subunits per asymmetric unit would give a reasonable V_m value of 2.87 ų/dalton. EcoRI endonuclease is the first protein which recognizes a specific sequence of bases in DNA to be crystallized in a form suitable for high resolution structure analysis.     

Crystallographic data of the selenoenzyme glutathione peroxidase

Glutathione peroxidase prepared from bovine erythrocytes yields small, but well-ordered plate-like crystals. X-ray investigation shows them to belong to monoclinic space group C2. Unit cell dimensions are: $\text{a = 90.4}\text{A}\text{?}\text{, b = 109.5}\text{A}\text{?}\text{, c = 58.6}\text{A}\text{?}$, β = 99 ° ± 15 min. The crystal density is ?_c = 1.36 ± 0.02 g.cm^?3. Consequently, the asymmetric unit of the crystal cell is occupied by one tetrameric molecule of M_r 84,000. Matthew's (1968) parameter Γ_M is calculated to be 1.71 ų/dalton.     

Visualization of drug-nucleic acid interactions at atomic resolution. IV. Structure of an aminoacridine--dinucleoside monophosphate crystalline complex, 9-aminoacridine--5-iodocytidylyl (3'--5') guanosine

9-Aminoacridine forms a crystalline complex with the dinucleoside monophosphate, 5-iodocytidylyl(3′–5′)guanosine (iodoCpG). These crystals are monoclinic, space group P2₁ with $\text{a = 13.98}\text{A}\text{?}\text{, b = 30.58}\text{A}\text{?}\text{, c = 22.47}\text{A}\text{?}$ and β = 113.9 °. The structure has been solved to atomic resolution by Patterson and Fourier methods, and refined by a combination of Fourier and sum-function Fourier methods. The asymmetric unit contains four 9-aminoacridine molecules, four iodoCpG molecules and 21 water molecules, a total of 245 atoms. 9-Aminoacridine demonstrates two different intercalative binding modes and, along with these, two slightly different intercalative geometries in this model system.The first of these is very nearly symmetric, the 9-amino group lying in the narrow groove of the intercalated base-paired nucleotide structure. The second shows grossly asymmetric binding to the dinucleotide, the 9-amino group lying in the wide groove of the structure. Associated with these two different intercalative binding modes is a difference in geometries in the structures. Although both structures demonstrate C3′ endo (3′–5′) C2′ endo mixed sugar puckering patterns (i.e. both cytidine residues have C3′ endo sugar conformations, while both guanosine residues have C2′ endo sugar conformations), with corresponding twist angles between base-pairs of about 10 °, they differ in the magnitude of the helical screw axis dislocation accompanying intercalation (Sobell et al., 1977a,b). In the pseudosymmetric intercalative structure, this value is about +0.5 Å, whereas in the asymmetric intercalative structure this value is about +2.7 Å. These conformational differences can be best described as a “sliding” of base-pairs on the intercalated acridine molecule.Although the pseudosymmetric intercalative structure can be used in 9-aminoacridine-DNA binding, the asymmetric intercalative structure cannot since this poses stereochemical difficulties in connecting neighboring sugar-phosphate chains to the intercalated dinucleotide. It is possible, however, that the asymmetric binding mode is related to the mechanism of 9-aminoacridine-induced frameshift mutagenesis (Sakore et al., 1977), and we discuss this possibility here in further detail.     

Crystallization and preliminary crystallographic investigation of a DNA-RNA hybrid

A complex between the complementary hexanucleotides ribo-C-A-A-A-A-G and deoxyribo-C-T-T-T-T-G has been crystallized. The space group was determined to be P2₁ with unit cell dimensions $\text{a = 26.9}\text{A}\text{?}\text{, b = 50.9}\text{A}\text{?}\text{, c = 62.0}\text{A}\text{?}\text{; α = γ = 90 °, β = 108 °}$. The crystals contain both the ribo- and the deoxyribo-hexanucleotide strands in a ratio of approximately 1:1. Each asymmetric unit contains the equivalent of four hybrid molecules.