Steven E. Ealick

Three-dimensional structure of recombinant human granulocyte-macrophage colony-stimulating factor.

The crystal structure of recombinant human granulocyte-macrophage colony-stimulating factor (rhGM-CSF) has been determined at 2.8 A resolution using multiple isomorphous replacement techniques. There are two molecules in the crystallographic asymmetric unit, which are related by an approximate non-crystallographic 2-fold axis. The overall structure is highly compact and globular with a predominantly hydrophobic core. The main structural feature of rhGM-CSF is a four alpha-helix bundle, which represents approximately 42% of the structure. The helices are arranged in a left-handed antiparallel bundle with two overhand connections. Within the connections is a two-stranded antiparallel beta-sheet. The tertiary structure of rhGM-CSF has a topology similar to that of porcine growth factor and interferon-beta. Most of the proposed critical regions for receptor binding are located on a continuous surface at one end of the molecule that includes the C terminus.

Structure of scorpion toxin variant-3 at 1·2 Å resolution

The crystal structure of the variant-3 protein neurotoxin from the scorpion Centruroides sculpturatus Ewing has been refined at 1.2 A resolution using restrained least-squares. The final model includes 492 non-hydrogen protein atoms, 453 protein hydrogen atoms, eight 2-methyl-2,4-pentanediol (MPD) solvent atoms, and 125 water oxygen atoms. The variant-3 protein model geometry deviates from ideal bond lengths by 0.024 A and from ideal angles by 3.6 degrees. The crystallographic R-factor for structure factors calculated from the final model is 0.192 for 17,706 unique reflections between 10.0 to 1.2 A. A comparison between the models of the initial 1.8 A and the 1.2 A refinement shows a new arrangement of the previously poorly defined residues 31 to 34. Multiple conformations are observed for four cysteine residues and an MPD oxygen atom. The electron density indicates that disulfide bonds between Cys12 and Cys65 and between Cys29 and Cys48 have two distinct side-chain conformations. A molecule of MPD bridges neighboring protein molecules in the crystal lattice, and both MPD enantiomers are present in the crystal. A total of 125 water molecules per molecule of protein are included in the final model with B-values ranging from 11 to 52 A2 and occupancies from unity down to 0.4. Comparisons between the 1.2 A and 1.8 A models, including the bound water structure and crystal packing contacts, are emphasized.

Architecture of scorpion neurotoxins: a class of membrane-binding proteins

Abstract The three-dimensional structure of a scorpion neurotoxin has been determined from high-resolution crystallographic data. The protein possesses a large flattened surface that contains many of the conserved residues and a high concentration of hydrophobic residues. It is likely that other scorpion toxins have this same overall structure, and that they bind to excitable membranes through sites on the conserved-hydrophobic surface of the molecule.

Structure-based design of inhibitors of purine nucleoside phosphorylase

Inhibitors of purine nucleoside phosphorylase may have therapeutic value in the treatment of T-cell proliferative diseases such as T-cell leukemia, in the suppression of host-versus-graft response in organ transplants, and in the treatment of T-cell-mediated autoimmune diseases. Competitive inhibitors of this enzyme have been designed using the three-dimensional structure of the enzyme determined by X-ray crystallography. This approach has resulted in the synthesis of the most potent and membrane-permeable inhibitors of purine nucleoside phosphorylase reported so far.

Inhibitors of purine nucleoside phosphorylase

Purine nucleoside phosphorylase (PNP) catalyzes reversible phosphorolysis of purine deoxy- and ribonucleosides with formation (d)Rib-1-P and corresponding bases. PNP plays a leading role in the cell metabolism of nucleosides and nucleotides, as well as in maintaining the immune status of an organism. The major aim of the majority of studies on the PNP is the detection of highly effective inhibitors of this enzyme, derivatives of purine nucleosides used in medicine as immunosuppressors, which are essential for creating selective T-cell immunodeficiency in a human body for organ and tissue transplantation. The present work is devoted to the study of the effects of some synthetic derivatives of purine nucleosides on activity of highly purified PNP from rabbit spleen and also from human healthy and tumor tissues of lung and kidneys. Purine nucleoside analogues modified at various positions of both the heterocyclic base and carbohydrate residues have been investigated. Several compounds, including 8-mercapto-acyclovir, 8-bromo-9-(3,4-hydroxybutyl)guanine, which demonstrated potent PNP inhibition, could be offered for subsequent study as immunosuppressors during organ and tissue transplantation.Disclosed is a compound containing a 2-amino-7-(R)-3H,5H-pyrrolo[3,2-d]pyrimidin-4-one wherein R is cyclohexenyl, cyclohexyl, or -CH2-R1, and wherein R1 is an optionally substituted heteroalicyclic, pyridinyl or alicyclic group. Also disclosed is a compound of formula (I), wherein R1 is H, NH?2?, or OCH3, R?2? is an optionally substituted cyclic group of 5-7 carbon atoms optionally containing one or more heteroatoms, R?3 and R4? are independently H or C?1?-4 alkyl, m is 0-4, n is 0-6, p is 0-1, X is CN, CSNH2, PO(OH)2, COOH, SO2NH2, NH2, OH CNHNH2, tetrazole, or triazole, COR?5? where R5 is C1-4 alkyl, CF3, NH2, or OC1-4 alkyl, and Y is O or NH.

X-ray crystallographic studies of three heterocyclic complexes of the type cis-Mo(CO)4{(R2PO)2Si(Me)R′} (R2 = OCH2CMe2CH2O; R′ = Me, But or R2 = Ph2; R′ = But)

Abstract X-Ray crystal structures for three complexes of the type cis -Mo(CO) 4 {(R 2 PO) 2 -Si(Me)R′} (I: R 2 = OCH 2 CMe 2 CH 2 O, R′ = Me; II: R 2 = OCH 2 CMe 2 CH 2 O, R′ = Bu t ; III: R 2 = Ph 2 , R′ = Bu t ) are reported. Complex I crystallizes in the monoclinic space group P 2 1 / n ( a 15.373(1), b 10.617(2), c 16.053(1) A; β 110.336(6)°; V 2460(7) A 3 ; Z = 4), complex II crystallizes in the triclinic space group P 1 ( a 9.6203(9), b 10.6899(15), c 16.0098(25) A; α 69.4716(2), β 68.8474(2), γ 66.7698(2)°; V 1368.7(3) A 3 ; Z = 2) and complex III crystallizes in the orthorhombic space group P 2 1 2 1 2 1 ( a 11.500(2), b 16.430(2), c 17.892(3) A; V 3380(2) A 3 ; Z = 4). The conformations of the chelate rings in these complexes are affected by the nature of the P and Si substituents. The conformations of the chelate rings in I, II and III are best described as a twist boat, a distorted chair and a chaise longue, respectively. In contrast, the conformations of the 1,3,2-dioxaphosphorinane rings in I and II do not appear to be affected by the nature of the Si substituents and are distorted chairs for both complexes. These studies indicate that the P and Si substituents have very different effects upon the steric and electronic properties of these ligands.

Crystallization and preliminary X-ray investigation of recombinant human interleukin 4☆

Crystals of recombinant human interleukin 4 have been grown from solutions of ammonium sulfate. The crystals are tetragonal, space-group P4(1)2(1)2 or P4(3)2(1)2; the unit cell axes are a = 92.2(1) A and c = 46.4(1) A. The crystals are stable to X-rays for at least three days and diffract beyond 2.8 A resolution. The crystals contain approximately 63% solvent, assuming there is one molecule in the asymmetric unit.