Alexander McPherson

Structure at 2.3 A resolution of the gene 5 product of bacteriophage fd: a DNA unwinding protein.

Abstract The structure of the gene 5 DNA unwinding protein from bacteriophage fd has been determined by X-ray diffraction analysis of single crystals to 2.3 A resolution using six isomorphous heavy-atom derivatives. The essentially globular monomer appears to consist of three secondary structural elements, a radically twisted three-stranded antiparallel β sheet and two distinct anti-parallel β loops, which are joined by short segments of extended polypeptide chain. The molecule contains no α-helix. A long groove, or arch, 30 A in length is formed by the underside of the twisted β sheet and one of the two β ribbons. We believe this groove to be the DNA binding region, and this is supported by the assignment of residues on its surface implicated in binding by solution studies. These residues include several aromatic amino acids which may intercalate or stack upon the bases of the DNA. Two monomers are maintained as a dimer by the very close interaction of symmetry related β ribbons about the molecular dyad. About six residues at the amino and carboxyl terminus are in extended conformation and both seem to exhibit some degree of disorder. The amimo-terminal methionine is the locus for binding the platinum heavy-atom derivatives and tyrosine 26 for attachment of the major iodine substituent.

Preliminary structure analysis of canavalin from jack bean.

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 R32 pseudo symmetry, P63 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 a staggered hexameric ring and related by perfect or near perfect 3 2 point group symmetry. The outside diameter of the molecule is approximately 65 A and there appears to be a large solvent channel coincident with the molecular triad.

Crystallization of a DNA-unwinding protein: Preliminary X-ray analysis of fd bacteriophage gene 5 product

Abstract The gene 5 protein from bacteriophage fd, which binds to single-stranded progeny fd DNA, was obtained as large single crystals and subjected to X-ray diffraction analysis. The crystals are of monoclinic space group C2 with a = 75.8 A , b = 28.0 A , c = 42.5 A and β = 103 °12′ . The unit cell has one molecule of 9800 daltons as the asymmetric unit.

Purification of mitogenic proteins from hura crepitans and Robinia pseudaccacia

Abstract The lectin-mitogens from Hura crepitans and Robinia pseudacaccia have been purified by affinity chromatography and compared to that from Abrus precatorius by sodium lauryl sulfate gel electrophoresis. Robinia lectin is quite similar to that from Abrus precatorius in that it consists of two distinct polypeptide chains of 32,000 and 30,000 daltons but unlike abrus lectin the chains are not joined by disulphide bonds. Hura lectin is composed of only a single polypeptide chain which migrates identically with the heavy chain of the abrus lectin. This heavy chain is likely responsible for binding to galactose residues on cell surfaces. The lectin from Robinia pseudaccacia has been obtained in crystalline form.

Crystallographic analysis of the phytoagglutinin from Abrus precatorius by X-ray diffraction and electron microscopy

Abstract The lectin from the seeds of Abrus precatorius has been crystallized and the crystals subjected to study by X-ray diffraction and electron microscopy. Three closely related crystal forms were obtained, of orthorhombic space group P 2 1 2 1 2 1 with a = 138 A , b = 142 A , and c = 173 A , of tetragonal space group P 4 1 2 1 2 with a = b = 136 A , c = 176 A , and a twinned intermediate of the first two. From electron microscopy and two-dimensional spatial filtering of electron micrographs of the crystals, the molecule appears to consist of four similar domains grouped in a roughly planar diamond-shaped arrangement having a local intramolecular dyad axis. The average diameter of the Abrus lectin molecule is 50 to 60 A and the individual domains appear to have a diameter of about 25 A.

Spatial filtering of electron micrographs of negatively stained α-amylase crystals

Abstract Images of the α-amylase molecule from pig pancreas have been obtained in two projections by the application of spatial filtering techniques to electron micrographs of negatively stained microcrystals of the enzyme. The Fourier filtering was performed on a PPD 11 40 digital computer after microdensitometry on an Optronics P-1000 photoscan system. The set of procedures and programs we employed are described herein. We compared these images with equivalent images obtained by three-dimensional X-ray diffraction analysis using conventional isomorphous replacement at approximately twice the resolution. We find that there is quite good agreement between the two kinds of images, and that a number of gross structural features compare quite well. We conclude that the combination of electron microscopy with digital spatial filtering when applied to protein microcrystals yields very respectable results.

Internal dihedral symmetry of α-amylase and α-mannosidase

Abstract Three-dimensional structure analyses of two polysaccharide degrading enzymes, α-amylase and α-mannosidase, though not yet complete, reveal that both enzymes are structural tandem duplicates and that the two similar domains within each are related by a dyad axis of symmetry.

Spatially filtered images of B. subtilisα‐amylase crystals

Microcrystals of B. subtilis α‐amylase have been negatively stained and examined by electron microscopy. The micrographs were then subjected to spatial filtering and averaging using a Fourier transform procedure programmed for a mini computer. From these ‘enhanced’ images refined crystallographic parameters were obtained and a model for the packing of the asymmetric units within the crystal was derived. In addition, these parameters obtained from electron microscopy are compared with those derived from limited X‐ray diffraction data on macrocrystals of the same form.

Preliminary X-ray diffraction data for chicken muscle glycerol 3-phosphate dehydrogenase

Abstract Single crystals of chicken muscle glycerol 3-phosphate dehydrogenase have been obtained which are suitable for a high resolution structure analysis. The crystals are triclinic with smallest cell dimensions of a=58.9 A b=54.5 A c=58.5 A α =91° β =95° δ =89° but show a substantial degree of pseudo symmetry which indicates the presence of at least an approximate and possibly exact dyad axis relating the two subunits in the asymmetric unit.