Crystallization of Phi29 spindle-shaped nano-bar anti-receptor with glycosidase domain.

Bacteriophage phi29 is a small, well-characterized dsDNA virus that infects Bacillus subtilis. The anti-receptor of phi29 consists of oligomers of the 854-residue protein gp12 and plays an essential role in infection initiation by binding to the receptor on the host cell surface. Oligomers of gp12 exhibit a narrow spindle-shaped configuration 15 nm in length as revealed by electron microscopy and thus are potentially useful nanoscale tools, building blocks, or motor arms. To understand the mechanism of viral infection initiation and to provide a basis for engineering recombinant gp12 for nanotechnology applications, we have initiated structural and bioinformatics studies of gp12. We report here the growth of crystals of gp12 that diffract to 3.0 A resolution. The space group is P3(1)21 or P3(2)21 with unit cell lengths of a = 84.4 A and c = 167.6 A. The asymmetric unit is predicted to contain one gp12 molecule and 32% solvent (VM = 1.8 A3/Da). Domain boundary analysis revealed that gp12 may harbor three domains besides a 24 residue auto-cleave region. The N-terminal half of gp12 contains a domain with about 400 residues that held 44% sequence identity to endopolygalacturonase, a fungal glycosyl hydrolase that catalyzes hydrolysis of the polygalacturonic acid alpha1-4 glycosidic linkage found in plant cell walls. Interestingly, the cell wall of Bacillus subtilis contains a polysaccharide component made from two sugar monomers, N-acetylmuramic acid and N-acetylglucosamine, which resemble alpha-galacturonic acid in that they possess a six-membered pyranose ring. Hence, polygalacturonic acid of plant cell walls and peptidoglycan of bacterial cell walls may offer a similar topography in relation to the polysaccharides. These results suggest a function for gp12 as a cell-wall degrading enzyme in addition to its role in recognition of the host receptor.

Slow vertical deposition of colloidal crystals: a Langmuir-Blodgett process?

For an evaporating colloidal suspension in which the evaporation velocity exceeds the sedimentation velocity, particles will accumulate at the solvent-air interface. If neither diffusion nor convection can disperse this accumulation, it is expected to grow into a colloidal multilayer several microns thick. We observe that the thickness of colloidal crystals vertically deposited from 1 mum diameter polystyrene latex suspensions of 0.002 < or = phi < or = 0.008 increases linearly with distance in the growth direction and that these thickness profiles are consistent with their growth from a horizontal colloidal layer accumulated beneath the solvent-air interface. We describe a means for performing vertical deposition at growth rates slower than the evaporation rate by adding solvent to the bottom of the colloidal suspension and observe that halving the growth rate of vertical deposition increases both the thickness and the reflectivity of the resulting colloidal crystals, effects indistinguishable from those of doubling the concentration of the colloidal suspension, data also consistent with the colloidal crystals' growth from a horizontal layer of particles beneath the interface. If sufficiently little reorganization is involved as particles move from this horizontal layer to the vertically deposited colloidal crystal, slow vertical deposition of polymer microspheres might be thought of as the Langmuir-Blodgett transfer of a horizontal colloidal crystal onto a vertical substrate. Colloidal crystals deposited using both high concentration and slowed growth can have peak IR reflectance in excess of 80%, exceeding most published values. These observations provide a conceptual framework for engineering vertically deposited colloidal crystals that combine thickness with good optical performance.