Crystal structure and ligand selectivity of the antineoplastic lectin from the common edible mushroom (Agaricus bisporus)

Crystal structure and ligand selectivity of the antineoplastic lectin from the common edible mushroom (Agaricus bisporus

Crystallization and preliminary X-ray study of the common edible mushroom (Agaricus bisporus) lectin

The lectin from the common edible mushroom Agaricus bisporus (ABL) belongs to the group of proteins that have the property of binding the Thomsen-Friedenreich antigen (T-antigen) selectively and with high affinity, but does not show any sequence similarity to the other proteins that share this property. The ABL sequence is instead similar to those of members of the saline-soluble fungal lectins, a protein family with pesticidal properties. The presence of different isoforms has been reported. It has been found that in order to be able to grow diffraction-quality crystals of the lectin, it is essential to separate the isoforms, which was performed by preparative isoelectric focusing. Using standard procedures, it was possible to crystallize the most basic of the forms by either vapour diffusion or equilibrium dialysis, but attempts to grow crystals of the other more acidic forms were unsuccessful. The ABL crystals belong to the orthorhombic space group C222(1), with unit-cell parameters a = 93.06, b = 98.16, c = 76.38 A, and diffract to a resolution of 2.2 A on a conventional source at room temperature. It is expected that the solution of this structure will yield further valuable information on the differences in the T-antigen-binding folds and will perhaps help to clarify the details of the ligand binding to the protein

A Phosphatidylinositol-Transfer Protein and Phosphatidylinositol-4-phosphate 5-Kinase Control Cdc42 to Regulate the Actin Cytoskeleton and Secretory Pathway in Yeast

The actin cytoskeleton rapidly depolarizes in yeast secretory (sec) mutants at restrictive temperatures. Thus, an unknown signal conferred upon secretion is necessary for actin polarity and exocytosis. Here, we show that a phosphatidylinositol (PI) transfer protein, Sfh5, and a phosphatidylinositol-4-phosphate 5-kinase, Mss4, facilitate Cdc42 activation to concomitantly regulate both actin and protein trafficking. Defects in Mss4 function led to actin depolarization, an inhibition of secretion, reduced levels of phosphatidylinositol 4,5-bisphosphate [PI(4,5)P2] in membranes, mislocalization of a pleckstrin homology domain fused to green fluorescent protein, and the mislocalization of Cdc42. Similar defects were observed in sec, myo2-66, and cdc42-6 mutants at elevated temperatures and were rescued by the overexpression of MSS4. Likewise, the overexpression of SFH5 or CDC42 could ameliorate these defects in many sec mutants, most notably in sec3Δ cells, indicating that Cdc42-mediated effects upon actin and secretion do not necessitate Sec3 function. Moreover, mutation of the residues involved in PI binding in Sfh5 led to the mislocalization and loss of function of both Sfh5 and Cdc42. Based upon these findings, we propose that the exocytic signal involves PI delivery to the PI kinases (i.e., Mss4) by Sfh5, generation of PI(4,5)P2, and PI(4,5)P2-dependent regulation of Cdc42 and the actin cytoskeleton