Association of the type II cAMP-dependent protein kinase with a human thyroid RII-anchoring protein. Cloning and characterization of the RII-binding domain.

The type II cAMP-dependent protein kinase (PKA) is localized to specific subcellular environments through binding of the dimeric regulatory subunit (RII) to anchoring proteins. Subcellular localization is likely to influence which substrates are most accessible to the catalytic subunit upon activation. We have previously shown that the RII-binding domains of four anchoring proteins contain sequences which exhibit a high probability of amphipathic helix formation (Carr, D. W., Stofko-Hahn, R. E., Fraser, I. D. C., Bishop, S. M., Acott, T. E., Brennan, R. G., and Scott J. D. (1991) J. Biol. Chem. 266, 14188-14192). In the present study we describe the cloning of a cDNA which encodes a 1015-amino acid segment of Ht 31. A synthetic peptide (Asp-Leu-Ile-Glu-Glu-Ala-Ala-Ser-Arg-Ile-Val-Asp-Ala-Val-Ile-Glu-Gln-Val -Lys-Ala-Ala-Tyr) representing residues 493-515 encompasses the minimum region of Ht 31 required for RII binding and blocks anchoring protein interaction with RII as detected by band-shift analysis. Structural analysis by circular dichroism suggests that this peptide can adopt an alpha-helical conformation. Both Ht 31 (493-515) peptide and its parent protein bind RII alpha or the type II PKA holoenzyme with high affinity. Equilibrium dialysis was used to calculate dissociation constants of 4.0 and 3.8 nM for Ht 31 peptide interaction with RII alpha and the type II PKA, respectively. A survey of nine different bovine tissues was conducted to identify RII binding proteins. Several bands were detected in each tissues using a 32P-RII overlay method. Addition of 0.4 microM Ht 31 (493-515) peptide to the reaction mixture blocked all RII binding. These data suggest that all anchoring proteins bind RII alpha at the same site as the Ht 31 peptide. The nanomolar affinity constant and the different patterns of RII-anchoring proteins in each tissue suggest that the type II alpha PKA holoenzyme may be specifically targeted to different locations in each type of cell.     

Serial manipulator functional calibration for in vitro biomechanical testing

Serial manipulators are often used in biomechanical testing of human joints because they are precise, repeatable instruments that can create interesting loading scenarios. Unfortunately, commercial serial manipulators often do not have acceptable global positional accuracy due to manufacturing tolerances, assembly errors, and other mechanical imperfections. Numerous calibration methods have been reported which calibrate geometric and non-geometric parameters to reduce static position errors under constant loading conditions. However, the manipulator's global accuracy during continuous motion with time-varying external loading conditions is often not addressed but is necessary for joint biomechanical testing. Using the Mitsubishi PA10-6CE as a case study, a novel functional calibration procedure was developed that performs both static and dynamic calibration. The calibration uses optimization techniques to populate a 34-parameter model that accounts for the robot's geometric and non-geometric parameters and significantly reduces the mean/peak static and dynamic position errors to 0.368/0.67 mm and 0.353/0.81 mm, respectively, while externally loaded.     

Modeling Interventions in the Owned Cat Population to Decrease Numbers,Knox County,TN

To find management strategies for controlling the owned cat population in Knox County, TN, the authors formulated a mathematical model using biological properties of such nonhuman animals and spay actions on certain age classes. They constructed this discrete-time model to predict the future owned cat population in this county and to evaluate intervention strategies to surgically sterilize some proportion of the population. Using the predicted population size and the number of surgeries for specific scenarios, they showed that focusing on specific age classes can be an effective feature in spay programs.     

The Effect of Space Flight on Monoclonal Antibody Synthesis in a Hybridoma Mouse Cell Line

The hybridoma cell line, 3G10G5, producing a monoclonal antibody to the major capsid protein VP1 from the avian polyomavirus budgerigar fledgling disease virus, was produced from a Balb/C mouse. This cell line was used to test the effects of microgravity on cellular processes, specifically protein synthesis. A time course study utilizing incorporation of [³⁵S]methionine into newly synthesized monoclonal antibody was performed on STS-77. After 5.5 days, it was observed that cell counts for the samples exposed to microgravity were lower than those of ground-based samples. However, radiolabel incorporation of the synthesized monoclonal antibody was similar in both orbiter and ground control samples. Overall, microgravity does not seem to have an effect on this cell line's ability to synthesize IgG protein.     

Metabolism of maltose and sucrose by microspores isolated from barley (Hordeum vulgare L.)

The aim of this work was to discover why barley (Hordeum vulgare L.) microspores die when cultured on media containing 40 mM sucrose but undergo embryogenesis on 40 mM maltose. Freshly isolated microspores were cultured for 6–24 h on media containing either [U-¹⁴C]maltose or [U-¹⁴C]sucrose at 40 mM, and the detailed distribution of ¹⁴C was determined. The amounts of glycolytic intermediates, ATP, ADP and AMP, in microspores were also measured. Cultures on sucrose differed from those on maltose in that the initial rate of metabolism was faster but declined rapidly, less ¹⁴C was recovered in polymers and more in alanine, there was extensive leakage of assimilated carbon, significant accumulation of ethanol and a lower adenylate energy charge. It is argued that microspores cultured on 40 mM sucrose die because they metabolize the sugar rapidly, become hypoxic and, as a result, accumulate large quantities of ethanol within the cells. Metabolism of maltose is slower and there is sufficient oxygen available to allow cells to survive in culture. Consequently some of the cultured cells undergo embryogenesis.P.S. thanks the Science and Engineering Research Council and Shell Research Ltd., Sittingbourne, for a Cooperative Award in Science and Engineering studentship.     

Repeated cadmium biosorption by regeneratedEnterobacter aerogenes biofilm attached to activated carbon

Summary The bacteriumEnterobacter aerogenes has been used to develop a biofilm over activated carbon for biosorption from various strength cadmium solutions (25–500ppm). High bacterial resistance to metal poisoning allowed biofilm regeneration to raise the net loading of cadmium over the carbon by repeated biosorption runs.     

Recent H3N2 Influenza Virus Clinical Isolates Rapidly Acquire Hemagglutinin or Neuraminidase Mutations When Propagated for Antigenic Analyses

Prior to serological testing, influenza viruses are typically propagated in eggs or cell culture. Recent human H3N2 strains bind to cells with low avidity. Here, we isolated nine primary H3N2 viral isolates from respiratory secretions of children. Upon propagation in vitro, five of these isolates acquired hemagglutinin or neuraminidase mutations that increased virus binding to cell surfaces. These mutations can potentially confound serological assays commonly used to identify antigenically novel influenza viruses.