Flipping a genetic switch by subunit exchange.

The bacteriophage T4 AsiA protein is a multifunctional protein that simultaneously acts as both a repressor and activator of gene expression during the phage life cycle. These dual roles with opposing transcriptional consequences are achieved by modification of the host RNA polymerase in which AsiA binds to conserved region 4 (SR4) of sigma(70), altering the pathway of promoter selection by the holoenzyme. The mechanism by which AsiA flips this genetic switch has now been revealed, in part, from the three-dimensional structure of AsiA and the elucidation of its interaction with SR4. The structure of AsiA is that of a novel homodimer in which each monomer is constructed as a seven-helix bundle arranged in four overlapping helix-loop-helix elements. Identification of the protein interfaces for both the AsiA homodimer and the AsiA-sigma(70) complex reveals that these interfaces are coincident. Thus, the AsiA interaction with sigma(70) necessitates that the AsiA homodimer dissociate to form an AsiA-SR4 heterodimer, exchanging one protein subunit for another to alter promoter choice by RNA polymerase.

X-ray crystallographic and analytical ultracentrifugation analyses of truncated and full-length yeast copper chaperones for SOD (LYS7): a dimer-dimer model of LYS7-SOD association and copper delivery.

Copper-zinc superoxide dismutase (CuZnSOD) acquires its catalytic copper ion through interaction with another polypeptide termed the copper chaperone for SOD. Here, we combine X-ray crystallographic and analytical ultracentrifugation methods to characterize rigorously both truncated and full-length forms of apo-LYS7, the yeast copper chaperone for SOD. The 1.55 A crystal structure of LYS7 domain 2 alone (L7D2) was determined by multiple-isomorphous replacement (MIR) methods. The monomeric structure reveals an eight-stranded Greek key beta-barrel similar to that found in yeast CuZnSOD, but it is substantially elongated at one end where the loop regions of the beta-barrel come together to bind a calcium ion. In agreement with the crystal structure, sedimentation velocity experiments indicate that L7D2 is monomeric in solution under all conditions and concentrations that were tested. In contrast, sedimentation velocity and sedimentation equilibrium experiments show that full-length apo-LYS7 exists in a monomer-dimer equilibrium under nonreducing conditions. This equilibrium is shifted toward the dimer by approximately 1 order of magnitude in the presence of phosphate anion. Although the basis for the specificity of the LYS7-SOD interaction as well as the exact mechanism of copper insertion into SOD is unknown, it has been suggested that a monomer of LYS7 and a monomer of SOD may associate to form a heterodimer via L7D2. The data presented here, however, taken together with previously published crystallographic and analytical gel filtration data on full-length LYS7, suggest an alternative model wherein a dimer of LYS7 interacts with a dimer of yeast CuZnSOD. The advantages of the dimer-dimer model over the heterodimer model are enumerated.

The differential expression of NGFS-like substance from fresh pilose antler of Cervus nippon Temminck.

Extracts of cartilage from antlers of non-mature deer, e.g. Cervus nippon Temminck, have been used as Chinese traditional medicines and restoratives. Since bioactivities attributed to the antler extracts resembled growth factor activities, the objective of our study was to compare the biological activities of different deer antler extracts with those of known growth factors. Extracts of the deer antlers were found to stimulate the growth of nerve fibers and to induce morphologic changes during differentiation and effect DNA synthesis in PC-12 cells, thus sharing with a known growth factor, NGF, several bioactivities.

Reduced productivity in adult yellowfever mosquito (Diptera: Culicidae) populations.

Male and female Aedes aegypti (L.) mosquitoes of the laboratory strain ROCK were irradiated with 130 mw of argon 514.5 nm laser microbeams for 0.04, 0.25, 0.4, and 0.5 s, respectively. Egg production, percentage hatch, and productivity (average number of adults surviving after 3 wk) were used to assess mutagenic effects. Mortality was high for males in all laser radiation groups and increased with time of exposure. Except for the group treated for 0.25 s, significant reductions in total F1 progeny also were demonstrated for all other experimentals when male parents were exposed to laser radiation. Females showed a high mortality when subjected to 0.4- and 0.5-s laser radiation. No F1 progeny were produced when parental females were exposed for 0.25, 0.4, and 0.5 s. Numbers of F1 progeny from females exposed to 0.04 s of laser radiation were significantly reduced. A comparison of weekly mean number of progeny showed that the important differences in productivity occurred during the first and second week, respectively, when either male or female adult parents were subjected to laser radiation.

Nutritional status and energy metabolism of crayfish (Procambarus clarkii, Girard) muscle and hepatopancreas.

1. Food deprivation resulted in significant decreases in muscle carbohydrate, lipid and water content and increased ATP, ADP, AMP and total adenylate levels over the 21-day experimental period. 2. In the hepatopancreas phosphoarginine was significantly higher on day 21 in the starved crayfish. 3. Muscle energy charges remained within optimal (unstressed) ranges, while hepatopancreatic energy charges of food-deprived crayfish fell into suboptimal (stressed) ranges, indicating the necessity of examining organs separately to accurately ascertain metabolic changes in response to stressors.

Energy metabolism in largemouth bass (Micropterus floridanus salmoides) from stressed and non-stressed environments: adaptations in the secondary stress response.

White muscle proteins, carbohydrates, lipids, adenylates, phosphagen and the AEC, (adenylate energy charge) were measured in bass inhabiting stressful and non-stressful environments. Within an environment, in June stressed bass had higher carbohydrates, while non-stressed bass had lower ATP, total adenylates and AEC. Comparisons between environments revealed: non-stressed bass had higher ATP/ADP and AEC's in May, CrP/ATP in June, and protein in July; while stressed bass had higher carbohydrate in June and lipid in July. Other metabolites varied insignificantly. AEC's of both groups were within the optimal range indicating physiological compensation (adaptation) of energy metabolism (secondary stress response) occurred in bass inhabiting the stressful environment.