Catalysis by dienelactone hydrolase: A variation on the protease mechanism

Dienelactone hydrolase (DLH), an enzyme from the β-ketoadipate pathway, catalyzes the hydrolysis of dienelactone to maleylacetate. Our inhibitor binding studies suggest that its substrate, dienelactone, is held in the active site by hydrophobic interactions around the lactone ring and by the ion pairs between its carboxylate and Arg-81 and Arg-206. Like the cysteine/serine proteases, DLH has a catalytic triad (Cys-123, His-202, Asp-171) and its mechanism probably involves the formation of covalently bound acyl intermediate via a tetrahedral intermediate. Unlike the proteases, DLH seems to protonate the incipient leaving group only after the collapse of the first tetrahedral intermediate, rendering DLH incapable of hydrolyzing amide analogues of its ester substrate. In addition, the triad His probably does not protonate the leaving group (enolate) or deprotonate the water for deacylation; rather, the enolate anion abstracts a proton from water and, in doing so, supplies the hydroxyl for deacylation. © 1993 Wiley-Liss, Inc.     

Estimation of Individual Growth Curves by Empirically Weighted Least Squares

The simultaneous estimation of individual growth curves and a mean growth curve is accomplished by weighted least squares. A polynomial curve is fitted for each individual and the polynomial parameters are linear functions of parameters corresponding to covariates. A simple, computationally efficient variance-covariance estimator is derived. The resultant estimate is used in the weighted least squares estimation. The results are compared to empirical Bayes estimation.     

Synthesis and in vitro bioactivity of C-terminal deleted analogs of human growth hormone-releasing factor

A series of C-terminal deleted analogs of human growth hormone-releasing factor (hGRF) with either an amidated or a free carboxylic acid C-terminus were synthesized by solid phase methodology. Their capacity to release growth hormone was tested on rat anterior pituitary cells in monolayer culture. A gradual decrease of bioactivity down to 23% relative to hGRF was noted when the C-terminal amino acids were deleted to hGRF (1-34)OH. Further deletions, however, did not decrease the bioactivity because the potencies of the fragments, hGRF(1-31)NH2, (1-30)NH2 and (1-29)NH2 remained at about 50% of that of hGRF. Continual deletion of residues to hGRF(1-23)NH2, (1-22)NH2 and (1-21)NH2 still yielded bioactive fragments with full intrinsic activity despite very low potency. Only with the deletion down to hGRF(1-19)NH2 did the bioactivity completely disappear. Thus, together with the data published in a previous paper (1), the minimal biologically active core of hGRF with full intrinsic activity comprises the fragment (3-21).     

Sex-specific somatic–otolith growth relationship in two Gadidae

Otolith and somatic mass of two Gadidae ( Merlangius merlangus and Trisopterus minutus ) were compared in order to analyse the sex-specific relationship between otolith and somatic growth at age. In the present study, otolith mass appeared a reliable indicator of age in both species. Otolith growth reflected somatic growth, but the relationship between these characters varied and differed between species and sexes.     

An Inhibitor of the UDP-N-Acetylgalactosamine:GM3, N-Acetylgalactosaminyltransferase: Purification and Properties, and Preparation of an Antibody to This Inhibitor

An inhibitor of the UDP-N-acetylgalactosamine:GM3, N-acetylgalactosaminyltransferase (EC 2.4.1.92) has been purified close to 100-fold from chicken blood serum. The method of purification includes heating, dialysis, passage through a column of DEAE-Sephadex, filtration through Amicon XM 100, and passage through Sepharose 6B. The molecular weight determined by Sepharose 6B was 200,000, but on sodium dodecyl sulfate-polyacrylamide gel electrophoresis it appears as if the compound dissociated into components of 68,000. The inhibitor was not active on other glycosyl transferases and lost its inhibitory activity following treatment with pronase and trypsin. alpha-Chymotrypsin did not affect the inhibitor. An antibody to this inhibitor was prepared which decreased its inhibitory capability and precipitated with it in a radial double immunodiffusion experiment.     

The effects of cell density and metabolite flux on cellular dynamics

Summary Density-dependent regulation of cell growth in tissue culture is a well-known phenomenon but the mechanism of regulation remains obscure. Here we explore the effects of cell density and metabolite flux on the collective dynamics of a cell population. The intracellular dynamics are modelled by positive feedback kinetic mechanisms of the kind known to apply to yeast cells. Several experimental observations related to glycolytic oscillations are predicted and it is suggested that the general conclusions may be applicable in a broader context.     

Ecology and biogeography of plant communities associated with the post Plio-Pleistocene relict Rhododendron ponticum subsp. baeticum in southern Spain

Aims Rhododendron ponticum L. is reputed to be a post Plio‐Pleistocene relict plant species with a disjunct distribution that comprises the Iberian Peninsula to the west and the Euxinian region plus some restricted Mediterranean areas to the east. We analysed the ecological range (of subsp. baeticum) in the western area (Aljibe Mountains, north of the Strait of Gibraltar) to understand the factors determining the present area limitation. Location Sierra del Aljibe, north of the Strait of Gibraltar (Iberian Peninsula). Methods We selected 20 riparian sites where R. ponticum is common, and compiled data on the ecological diversity of associated woody species and ferns. We established a 500‐m main transect in each site, along the stream or river course, in which we placed five 20‐m‐long plots at regular intervals. We recorded physiographic habitat features, woody plants and fern abundance, and the number of R. ponticum individuals. Results Rhododendron ponticum in southern Spain is restricted to riparian forests in acidic soils (pH 4.0–6.4), and is mainly found on the banks of inclined and enclosed streams. In our inventory we recorded 59 woody taxa and 12 ferns, with R. ponticum being the dominant species of the understorey (mean abundance 78.6%). The communities are characterized by a high incidence of the humid warm temperate element, both in number of species (18.8 ± 3.7 per site) and abundance; meanwhile, the presence of the modern Mediterranean element (mean number of species 3.4 ± 3.8 per site) appears to be favoured by disturbance. These ecological–historical groups of taxa also show distinct patterns of typological habit, frequency of endemism, infrageneric diversity and geographical range. Populations of R. ponticum are characterized by a very variable density of seedlings in many sites, and the virtual lack of juveniles. Main conclusions Riparian forests of the Aljibe Mountains constitute a refuge for R. ponticum where the species persists, but populations appear to be in decline. The narrow ecological range of R. ponticum in the area strongly contrasts with its wide amplitude in the eastern natural area, mainly the Euxinian region, where R. ponticum probably finds better conditions due to the environmental heterogeneity of the region, and the lack of a hot dry season.