Methane fluxes from differentially managed grassland study plots: the important role of CH4 oxidation in grassland with a high potential for CH4 production.

Methane oxidation fluxes were monitored with the closed chamber method in eight treatment plots on a semi-wet grassland site near Giessen, Germany. The management regimes differed in the amount of nitrogen (NH4NO3) fertilizer applied and in the height of the in-ground water table. No inhibition of CH4 oxidation occurred, regardless of the amount of annual N fertilizer applied. Instead, the mean CH4 consumption rates were correlated with the mean soil moisture of the plots. However, the correlation between daily soil water content and corresponding CH4 oxidation rate was always weak. During drought period (late summer) water stress was observed to restrict CH4 oxidation rates. The findings led to the question whether methane production with soil depth might modify the CH4 fluxes measured at the surface. Therefore, two new methods were applied: (1) soil air sampling with silicone probes; and (2) anaerobic incubations of soil cores to test for the methane production potential of the grassland soil. The probe measurements revealed that the CH4 sink capacity of a specific site was related to the vertical length of its CH4 oxidizing column, i.e. the depth of the CH4 producing horizon. Anaerobically incubated soil cores produced large amounts of CH4 comparable with tropical rice paddy soil. Under field conditions, heavy autumnal rain in 1998 led to a dramatic increase of soil CH4 concentrations upto 51 microliters l-1 at a depth of 5 cm. Nevertheless, no CH4 was released when soil surface CH4 fluxes were measured simultaneously. The results thus demonstrate the high CH4 oxidation potential of the thin aerobic topsoil horizon in a non-aquatic ecosystem.

Cleavage of recombinant and cell derived human immunodeficiency virus 1 (HIV-1) Nef protein by HIV-1 protease.

Recombinant purified Nef protein of HIV-1, as well as Nef protein derived from extracts of permanently HIV-1 infected glioblastoma cells and monocytes, are specifically cleaved by the HIV-1 protease. Nef cleavage products in cellular extracts treated with protease showed identical molecular weights as those obtained by digestion of purified Nef with recombinant HIV-1 protease. Since cellular extracts were prepared by detergent and mechanical lysis it cannot be excluded that physiological cytoplasmic conditions were altered. The lack of Nef cleavage by endogenous HIV-1 protease in infected cells might be due to low concentrations of viral protease and the presence of Gag precursor molecules as natural substrate. Using a panel of monoclonal antibodies two cleavage fragments of 19 kDa and 8 kDa were defined. The cleavage site was located by microsequencing between amino acid 57 and 58 (AW*LEAQEEEEVGF). The conserved cleavage motif within HIV-1 Nef suggests a potential biological function of Nef processing.

Identification of Mycelium-Associated Cellulase from Streptomyces reticuli.

Among 180 Streptomyces strains tested, 25 were capable of hydrolyzing microcrystalline cellulose (Avicel) at 30 degrees C. Streptomyces reticuli was selected for further studies because of its ability to grow at between 30 and 50 degrees C on Avicel. Enzymatic activities degrading Avicel, carboxymethyl cellulose, and cellobiose were found both in the culture supernatant and in association with the mycelium and crystalline substrate. The bound enzymes were efficiently solubilized by repeated washes with buffer of low ionic strength (50 mM Tris hydrochloride [pH 7.5]) and further purified by fast protein liquid chromatography. A high-molecular-weight Avicelase of >300 kilodaltons could be separated from carboxymethyl cellulase (CMCase) and beta-glucosidase activities (molecular mass, 40 to 50 kilodaltons) by gel filtration on Superose 12. The CMCase fraction was resolved by Mono Q anion-exchange chromatography into two enzymes designated CMCase 1 and CMCase 2. The beta-glucosidase activity was found to copurify with CMCase 2. The purified cellulase components showed optimal activity at around pH 7.0 and temperatures of between 45 and 50 degrees C. Avicelase (but not CMCase) activity was stimulated significantly by the addition of CaCl(2).