A new approach for calculating association constants from macromolecule-ligand binding isotherms

The calculation of association constants from measurements of the average number of ligands bound to a macromolecule can be reduced to the solution of a set of linear algebraic equations. However, in practice various approximations and graphical procedures have been preferred because the equations are very ill conditioned. A method is proposed in which a formal integration transforms the equations into the Vandermonde form, for which special methods of solution are available. The required integration is carried out by spline functions in an iterative procedure. The method gave satisfactory results for synthetic data based on the association constants for the binding of copper ions to ovalbumin, provided the data were extended to very low concentrations. Also a common error in the use of the extended Scatchard method for determining association constants is analyzed.     

An Automated Technique for the Radiographic Determination of Bone Age

We have developed an automated procedure for determining bone-age in the monkey, Macaca fascicularis. The status of epiphysial junctions and sesamoids were determined from radiographs. Epiphyses were scored as either open or closed, and sesamoids were scored as either present or absent. Bone-age was calculated by a computer program that stored and used a data base of probabilities for epiphysial closure and sesamoid appearance. The simplicity of scoring, combined with the speed and accuracy of the calculations, make this technique potentially adaptable for use in a variety of primate species.     

Photosynthetic responses of Zostera marina L. (Eelgrass) to in situ manipulations of light intensity

Summary Photosynthetic responses of the temperate seagrass, Zostera marina L., were examined by manipulations of photon flux density in an eelgrass bed in Great Harbor, Woods Hole, MA during August 1981. Sun reflectors and light shading screens were placed at shallow (1.3 m) and deep (5.5 m) stations in the eelgrass bed to increase (+35% to +40%) and decrease (-55%) ambient photon flux densities. The portion of the day that light intensities exceeding the light compensation point for Z. marina (H _comp) and the light saturation point (H _sat) were determined to assess the impact of the reflectors and shades. The H _comp and H _sat periods at the deep station shading screen were most strongly affected; H _comp was reduced by 11% and H _sat was reduced by 52%. Light-saturated photosynthetic rates, dark respiration rates, leaf chlorophyll content, chlorophyll a/b, PSU_{O 2} size, PSU density, leaf area, specific leaf area, leaf turnover times and leaf production rates were determined at the end of three sets of 1- to 2-week experiments. None of the measured parameters were affected by the photon flux density manipulations at the shallow station; however, at the deep station leaf production rates were significantly reduced under the shading screen and chlorophyll a/b ratios were higher at the reflector. These results indicate that adjustment to short-term changes in light regime in Z. marina is largely by leaf production rates. Further, the most dramatic changes in the periods of compensating or saturating photon flux densities had the greatest impact on the measured photosynthetic responses.     

Muramic Acid Assay in Sediments

An improved chromatographic assay for muramic acid which is sufficiently sensitive for marine sandy sediments is described; it involves acid hydrolysis, thin-layer chromatography, and gas-liquid chromatography.     

The size of the extracellular space in the isolated midgut ofManduca sexta

The size of the extracellular space in the isolated midgut ofManduca sexta measured statically with labeled sucrose and with labeled inulin was 48 and 50 percent of tissue water respectively. Both labeled sulfate and labeled mannitol yielded loading spaces which approached the volume of the tissue water with increasing pulse time and are not valid markers of the extracellular space. A washout method yielded unreliable values for the sucrose and inulin spaces and its use cannot be justified using presently available midgut chamber designs. Values of the sucrose space measured statistically in this study and other studies using different Lepidopteran larvae and different perfusion chambers range from 42 to 48 percent of the tissue water. Agreement between published values of the sulfate space and these values of the sucrose space is fortuitous, the labeling pulses being short enough to yield approximately half saturation of the actual sulfate loading space.     

Microspatial heterogeneity in the distribution of ciliates in a small pond

Five transects of contiguous samples from the surface of a small pond and one transect from its bottom were collected in order to quantify microspatial heterogeneity in the distribution of ciliated protozoa. Examination of the frequency-abundance relations for these transects suggests that they can be approximated by negative binomial distributions with a commonk of 1.87. Contagiousness or crowding increases with population density.Mean patch size and mean interpatch distance were measured for 4 transects as 1.5 to 2 cm and 3 to 4 cm, respectively. This heterogeneity is suggested to arise from behavioral aggregation about discrete food sources and be very ephemeral.Blocking of adjacent contiguous samples was used to investigate the effect of sample size on the apparent correlation between the numbers of pairs of taxa. In all cases examined, taxa were relatively independent in their distribution at small sample sizes and became more negatively or positively associated as samples were combined. This may reflect that the small scale patches are essentially monospecific.     

A steady-state kinetic study on the catalytic mechanism of ribulose bisphosphate carboxylase from soybean

The catalytic mechanism of soybean ribulose bisphosphate carboxylase was examined, through a study of the steady-state kinetic behavior of the fully activated enzyme using short time assays. The effects of substrates, products, alternative products, and two dead-end inhibitors were investigated. High concentrations of both substrates were observed to lead to nonhyperbolic relationships: concentrations of bicarbonate greater than 15 mm inhibited and concentrations of ribulose bisphosphate greater than 0.2–0.5 mm stimulated enzyme activity over that expected from a hyperbolic fit to the data. The kinetic patterns obtained and the nonhyperbolic behavior of substrates are interpreted to suggest that the binding of substrates and the release of products follow a steady-state random mechanism. The substrate activation by ribulose bisphosphate is likely to be physiologically significant.     

Frost-weathering on Mars: Experimental evidence for peroxide formation

Summary A laboratory study of the interaction of H₂O frost with samples of the minerals olivine (Mg,Fe)₂SiO₄ and pyroxene (Mg,Fe)SiO₃ at –11°C to –22°C revealed that an acidic oxidant was produced. Exposure of the frost-treated minerals to liquid H₂O produced a sudden drop in pH and resulted in the production of copious O_2(g) (as much as ~ 10²⁰ molecules g^–1). Exposure of frost-treated samples to 5 ml of 0.1M HCOONa solution resulted in the rapid oxidation of up to 43% of the formate to CO_2(g). These reactions were qualitatively similar to the chemical activity observed during the active cycles of the Viking lander Gas Exchange and Labeled Release Biology experiments. Attempts to identify the oxidant by chemical indicators were inconclusive, but they tentatively suggested that chemisorbed hydrogen peroxide may have formed. The formation of chemisorbed peroxide could be explained as a byproduct of the chemical reduction of the mineral. The following model was proposed. H^c was incorporated into the mineral from surface frost. This would have left behind a residual of excess OH^– _(ads) (relative to surface H⁺). Electrons were then stripped from the surface OH^– _(ads) (due to the large repulsive potential between neighboring OH^– _(ads)) and incorporated into the crystal to restore charge balance and produce a chemical reduction of the mineral. The resultant surface hydroxyl radicals could then have combined to form the more stable chemisorbed hydrogen peroxide species. While the chemisorbed peroxide should be relatively stable at low temperatures, it should tend to decay to O_(ads) + H₂O_(g) at higher temperatures with an activation energy of 34 kcal mole^–1. This is consistent with the long-term storage and sterilization behavior of the Viking soil oxidants. It is possible that as little as 0.1–1% frost-weathered material in the Martian soil could have produced the unusual chemical activity that occurred during the Viking Gas Exchange and Labeled Release experiments.This paper contains the material given in invited presentations at the COSPAR Meeting, Innsbruck, Austria, 5–7 June 1978 and at the Second Conference on Simulation of Mars Surface Properties, NASA Ames Research Center, 17–18 August 1978