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W Minor

Publications and source records attributed to W Minor.

23 records · Page 2Linked to original sources
PubMed1993

Crystallographic determination of the active site iron and its ligands in soybean lipoxygenase L-1.

Five ligands of the active site iron atom in soybean lipoxygenase L-1 have been identified from the electron density map of the crystallized enzyme. The position of the iron atom can be readily and independently located from an anomalous difference electron density map. The ligands identified are His-499, His-504, His-690, Asn-694, and Ile-839, the carboxy-terminal residue. Our previous view that these three histidines are essential for activity and binding of iron, based on site-specific mutation studies, is confirmed. A sixth protein ligand is not present, and the sixth coordination site opens into a wide cleft. The structure of the soybean lipoxygenase was solved by multiple anomalous isomorphous replacements.

Amino Acid Sequence↗
PubMed1992

The structure determination of Sindbis virus core protein using isomorphous replacement and molecular replacement averaging between two crystal forms.

The structure of Sindbis virus core protein has been determined by a combination of multiple isomorphous replacement and molecular replacement averaging techniques. The multiple isomorphous replacement phase determinations were made for two crystal forms (P2(1) and P4(3)2(1)2) of the core protein. The real-space molecular replacement averaging was subsequently carried out between two copies of the protein per asymmetric unit in the monoclinic form and one copy in the tetragonal form. This greatly improved the quality of the electron density maps. The Sindbis virus core protein polypeptide could be traced and related to the known amino acid sequence. The averaging procedure between different crystal forms, as described in this paper, should be generally applicable to other systems.

Crystallization↗
PubMed1991

Structure of Sindbis virus core protein reveals a chymotrypsin-like serine proteinase and the organization of the virion.

Sindbis virus consists of a nucleocapsid core surrounded by a lipid membrane through which penetrate 80 glycoprotein trimers. The structure of the core protein comprising the coat surrounding the genomic RNA has been determined. The polypeptide fold from residue 114 to residue 264 is homologous to that of chymotrypsin-like serine proteinases with catalytic residues His 141, Asp 163 and Ser 215 of the core protein positioned as in other serine proteinases. The C-terminal tryptophan remains in the P1 substrate site subsequent to the autocatalytic cis cleavage of the capsid protein, thus rendering the proteinase inactive. Model building of the Sindbis core protein dimer shows that the nucleocapsid is likely to have T = 4 quasisymmetry.

Amino Acid Sequence↗
PubMed1991

Removal of salt from a salt-induced protein crystal without cross-linking. Preliminary examination of "desalted" crystals of phosphoglucomutase by X-ray crystallography at low temperature.

A model procedure for removing salt from relatively fragile salt-induced protein crystals is proposed. The procedure is based on physical principles and is validated by using millimeter-size crystals of rabbit muscle phosphoglucomutase grown from a 2.1 M solution of ammonium sulfate. Three types of operations are included in the procedure: initial transfer to salt solutions of reduced concentration; transfer to the organic-rich phase of an equilibrium biphasic mixture obtained with aqueous solutions of polyoxyethylene and the salt; and addition of various replacement cosolutes in aqueous solutions of polyoxyethylene to reduce osmotic stress on the crystal as the remaining salt is removed. A critical feature of the overall procedure is maintenance of near equilibrium throughout by using a large number of steps involving small changes in solute concentration. The conditions used in the actual transfer were adjusted to eliminate the fracturing of crystals by visually distinguishing between two opposing types of fracture patterns: those produced by osmotic crushing as opposed to osmotic expansion. Basic requirements for a successful procedure with other protein crystals are a high permeability toward small solutes and a relatively slow dissolution rate at salt concentrations for which biphasic mixtures can be obtained. Desalted crystals of phosphoglucomutase have no visible fractures, are stable in the final solution for at least a week, and exhibit no noticeable change in the resolution of their X-ray diffraction pattern. In fact, desalted crystals can be rapidly cooled to 160 K, whereas untreated crystals are almost completely disordered by the same cooling procedure. The component of the desalting mixture whose presence is crucial to the success of the cooling process is polyoxyethylene, which apparently impedes the formation of ice within the protein crystal. Diffraction data obtained with an area-detector diffractometer did not differ significantly, either in terms of quality or resolution range, between crystals in 2.3 M ammonium sulfate at room temperature and crystals at 160 K in which ammonium sulfate had been replaced by glycine. The successful use of the following replacement solutes, instead of glycine, also is documented: sucrose, glycerol, and a low molecular weight poly(ethylene glycol) (PEG-400).

Animals↗