Enhancin, the granulosis virus protein that facilitates nucleopolyhedrovirus (NPV) infections, is a metalloprotease.

Enhancin is a Trichoplusia ni granulovirus protein that facilitates nucleopolyhedrovirus (NPV) infections in lepidopterans. Gel filtration and ion exchange chromatography and immobilized alpha-macroglobulin were used to purify this protein and the removal of the contaminating proteases did not diminish the in vivo or in vitro activity of enhancin. Metal chelators were the only protease inhibitors capable of preventing digestion of the peritrophic membrane (PM) proteins by enhancin, indicating that enhancin is a metalloprotease. In addition, the canonical zinc binding site, HEXXH, found in most metalloproteases, was identified in the sequences of enhancins from three different granuloviruses. The identity of enhancin as a metalloprotease that facilitates NPV infections in lepidopterous larvae was confirmed by the expression of enhancin in a recombinant Autographa californica MNPV-baculovirus system and the purification of a recombinant enhancin that was active in neonate bioassays and that digested specific PM proteins. The recombinant enhancin was also inhibited by metal chelators and both the native and recombinant enhancin could be reactivated by divalent ions, further confirming that enhancin is a metalloprotease.

Comparison of the lipid acyl chain dynamics between small and large unilamellar vesicles.

13C NMR spin-lattice relaxation (T1) rates and 13C-1H nuclear Overhauser effects (NOEs) were measured in an identical fashion in two lipid preparations having dramatically different curvatures. The T1 times that were obtained at four magnetic field strengths were fit along with the NOEs to simple models for lipid molecular dynamics. The results indicate that phospholipid chain ordering and dynamics are virtually identical in small and large unilamellar vesicles at the time scales sampled by these 13C-NMR studies. The order parameters and reorientational correlation times that characterize the amplitudes and rates of internal acyl chain motions were equal within experimental error for the methylene segments in the middle of the chains. The only significant differences in order parameters and correlation times between the two vesicle types were small and appeared at the ends of the acyl chains. At the carbonyl end the order was slightly higher in small vesicles than large vesicles, and at the methyl end the order was slightly lower for small vesicles. This indicates that in the more planar systems the acyl chains exhibit a slightly flatter order profile than in more highly curved membranes. The use of the same experimental approach in both small and large vesicle systems provided a more reliable and accurate assessment of the effect of curvature on molecular order than has been previously obtained.