INORGANIC CARBON LIMITATION OF PHOTOSYNTHESIS IN ULVA ROTUNDATA (CHLOROPHYTA)1

A computerized oxygen electrode Astern was used to make rapid and accurate measurements of photosynthetic light and dissolved inorganic carbon (DIC) response cures with a macroalga. Ulva rotundata Blid. was grown in an outdoor, continuous flow system in seawater under sunlight or 9% of sunlight at Beaufort, North Carolina. The light compensation points in the shade- and sun-grown plants, measured in seawater, were at photon flux densities (PFDs) of 16 and 27 μmol. Photons·m^?2·s^?1, respectively but the quantum yield of O₂ evolution was not significantly different. Rates of photosynthesis in seawater per unit area of thallus under saturating light and rates of dark respiration were about 1.5-fold higher in sun- than in shade-grown plants. The concentration of DIC in seawater (approximately 2 mM) limited photosynthesis at absorbed PFDs above 60–70 μmol photons·m^?2·s^?1 Addition of 20 mM inorganic carbon had no effect on quantum yield but caused about a 1.5-fold increase in the light-saturated photosynthetic rate in both shade- and sun-grown Ulva. The effect of DIC supplementation was greatest in plants grown in October and least in plants grown in June. The light- and DIC-saturated rate of photosynthesis in seawater was similar to the maximum rate obtained by exposing Ulva to 10% CO₂, in the gas phase. The carbon isotope values (δ¹³C, reflecting the ¹³C/¹²C ratio compared to a standard) of Ulva grown in the same seawater supply were dependent on light and agitation. Samples from Beaufort Inlet were more negative (δ¹³C value, ?20.03‰) than those grown in bright light with agitation (δ¹³C value, ?17.78‰ outdoors; ?17.23‰ indoors), which may indicate DIC supply limited carbon uptake in seawater.     

Holocene vegetation and water level history in two bogs of the Cordillera de Talamanca, Costa Rica

Pollen records of Holocene sediment cores from the Costa Rican Cordillera de Talamanca (La Chonta bog, 2310 m and La Trinidad bog, 2700 m) show the postglacial development of the montane oak forest zone from ca. 9500 to 1500 yr BP. During the early Holocene (ca. 9500–700 yr BP), alder vegetation covered the La Chonta and La Trinidad bogs and their adjacent hills. The upper forest line is inferred to be at 2800–3000 m elevation. A Podocarpus-Quercus forest characterised the middle Holocene (ca. 7000–4500 yr BP). The upper forest line is located at >3000 m reaching the present-day altitudinal distribution. A Quercus forest characterised the late Holocene (ca. 4500–1500 yr BP). Compared to modern conditions, the early Holocene has similar average temperatures, but the moisture level was probably higher. Pollen evidence for the late Holocene indicates drier environmental conditions than today. In order to improve the paleoecological interpretation, we described the local vegetation and used moss samples as pollen traps at both montane bogs along strong soil moisture gradients.The Netherlands Centre for Geo-ecological Research, ICG     

Body Position,Age and Mass Effect of Adiposity on Adipose Tissue Red Cell Flux in Morbid Obesity

Objective: To measure red cell flux of adipose tissue in morbidly obese patients' pannus in the upright and supine position to determine factors which would render the lower pannus susceptible to ischemic necrosis. Design: A cohort study of morbidly obese subjects without ischemic necrosis. Setting: University teaching hospital. Patients: Twenty-three consecutive morbidly obese patients referred for gastroplasty. Measurements: Red cell flux, measured as RMS voltage by a laser Doppler velocimeter. An optical fiber with a tip diameter of 250μ was inserted into the upper and lower pannus and output recorded in the upright and supine positions. Other variables recorded were age, BMI, blood pressure and serum lipids. Results: Adipose tissue red cell flux demonstrates considerable spatial and temporal heterogeneity from subject to subject and in various locations in the pannus. No differences in red cell flux were detected in response to change in position. However, regression analysis demonstrated that the gradient between the upper and lower abdomen in the supine position was increasingly positive with age and in the upright position it was increasingly positive with increasing weight or BMI. Conclusions: These data suggest that red cell flux is heterogeneously distributed in the abdominal pannus and is not greatly influenced by body position. However, with increasing age and adiposity there is a gradient for decreased red cell flux to the lower portion of the pannus. This may be a factor in rendering this part of the pannus prone to ischemic fat necrosis.     

A carbon monoxide irreducible form of cytochrome c oxidase and other unusual properties of the “monomeric” shark enzyme

Contrary to previous reports, the functional and spectral properties of “monomeric” shark cytochrome c oxidases are not entirely similar to those of the “dimeric” beef enzyme. Most significantly, unlike the behavior of beef oxidase, the fully oxidized shark enzyme is not reducible by carbon monoxide. Also, preparations of the shark enzyme, isolated at pH 7.8-8.0, lead to more than 60% of the sample always being obtained in a resting form, whereas similarly prepared beef oxidase is very often obtained, both by ourselves and others, exclusively in the pulsed form. Although the electronic absorption, magnetic circular dichroism and electron paramagnetic resonance (EPR) spectra of cytochrome c oxidase obtained from several shark species are similar to those of the beef enzyme, there are some significant differences. In particular, the Soret maximum is at 422 nm in the case of the fully oxidized resting shark oxidases at physiological pH and not 418 nm as commonly found for the beef enzyme. Moreover, the resting shark oxidases do not necessarily exhibit a “g = 12” signal in their EPR spectra. The turnover numbers of recent preparations of the shark enzyme are higher than previously reported and, interestingly, do not differ within experimental uncertainty from those documented for several beef isoenzymes assayed under comparable conditions.     

Self-Incorporation of coenzymes by ribozymes

RNA molecules that are assembled from the four standard nucleotides contain a limited number of chemical functional groups, a characteristic that is generally thought to restrict the potential for catalysis by ribozymes. Although polypeptides carry a wider range of functional groups, many contemporary protein-based enzymes employ coenzymes to augment their capabilities. The coenzymes possess additional chemical moieties that can participate directly in catalysis and thereby enhance catalytic function. In this work, we demonstrate a mechanism by which ribozymes can supplement their limited repertoire of functional groups through RNA-catalyzed incorporation of various coenzymes and coenzyme analogues. The group I ribozyme of Tetrahymena thermophila normally mediates a phosphoester transfer reaction that results in the covalent attachment of guanosine to the ribozyme. Here, a shortened version of the ribozyme is shown to catalyze the self-incorporation of coenzymes and coenzyme analogues, such as NAD⁺ and dephosphorylated CoA-SH. Similar ribozyme activities may have played an important role in the RNA world, when RNA enzymes are thought to have maintained a complex metabolism in the absence of proteins and would have benefited from the inclusion of additional functional groups.Correspondence to: G.F. Joyce     

Response regulators of bacterial signal transduction systems: Selective domain shuffling during evolution

Response regulators of bacterial sensory transduction systems generally consist of receiver module domains covalently linked to effector domains. The effector domains include DNA binding and/or catalytic units that are regulated by sensor kinase-catalyzed aspartyl phosphorylation within their receiver modules. Most receiver modules are associated with three distinct families of DNA binding domains, but some are associated with other types of DNA binding domains, with methylated chemotaxis protein (MCP) demethylases, or with sensor kinases. A few exist as independent entities which regulate their target systems by noncovalent interactions.In this study the molecular phylogenies of the receiver modules and effector domains of 49 fully sequenced response regulators and their homologues were determined. The three major, evolutionarily distinct, DNA binding domains found in response regulators were evaluated for their phylogenetic relatedness, and the phylogenetic trees obtained for these domains were compared with those for the receiver modules. Members of one family (family 1) of DNA binding domains are linked to large ATPase domains which usually function cooperatively in the activation of E. Coli ⁵⁴-dependent promoters or their equivalents in other bacteria. Members of a second family (family 2) always function in conjunction with the E. Coli ⁷⁰ or its equivalent in other bacteria. A third family of DNA binding domains (family 3) functions by an uncharacterized mechanism involving more than one a factor. These three domain families utilize distinct helix-turn-helix motifs for DNA binding.The phylogenetic tree of the receiver modules revealed three major and several minor clusters of these domains. The three major receiver module clusters (clusters 1, 2, and 3) generally function with the three major families of DNA binding domains (families 1, 2, and 3, respectively) to comprise three classes of response regulators (classes 1, 2, and 3), although several exceptions exist. The minor clusters of receiver modules were usually, but not always, associated with other types of effector domains. Finally, several receiver modules did not fit into a cluster. It was concluded that receiver modules usually diverged from common ancestral protein domains together with the corresponding effector domains, although domain shuffling, due to intragenic splicing and fusion, must have occurred during the evolution of some of these proteins.Multiple sequence alignments of the 49 receiver modules and their various types of effector domains, together with other homologous domains, allowed definition of regions of striking sequence similarity and degrees of conservation of specific residues. Sequence data were correlated with structure/function when such information was available. These studies should provide guides for extrapolation of results obtained with one response regulator to others as well as for the design of future structure/function analyses. Correspondence to: M.H. Saier, Jr.     

The measurement of transmembrane helices by the deconvolution of CD spectra of membrane proteins: A review

The interpretation of the CD spectra of proteins to date requires additional secondary structural information of the proteins to be analyzed, such as x-ray or nmr data. Therefore, these methods are inappropriate for a CD data base whose secondary structures are unknown, as in the case of the membrane proteins. The Convex Constraint Analysis algorithm [A. Perczel, M. Hollósi, G. Tusnády, and G. D. Fasman (1991) Protein Engineering, Vol. 4, 669–679], on the other hand, operates only on a collection of spectral data to extract the common spectral components with their spectral weights. The linear combinations of these derived “pure” CD curves can reconstruct the original data set with great accuracy. For a membrane protein data set, the five-component spectra so obtained from the deconvolution consisted of two different types of α-helices (the α-helix in the soluble domain and the α_T-helix, for the transmembrane α-helix), a β-pleated sheet, a class C-like spectrum related to β-turns, and a spectrum correlated with the unordered conformation. The deconvoluted CD spectrum for the α_T-helix was characterized by a positive red-shifted band in the range 195–200 nm (+95,000 deg cm² dmol^?l), with the intensity of the negative band at 208 nm being slightly less negative than that of the 222 nm band (?50,000 and ?60,000 deg cm² dmol^?1, respectively) in comparison with the regular α-helix, with a positive band at 190 nm and two negative bands at 208 and 222 nm with magnitudes of + 70,000, ?30,000, and ?30,000 deg cm² dmol^?1, respectively. © 1994 John Wiley & Sons, Inc.     

Upregulation of interferon-induced indoleamine 2,3-dioxygenase in human macrophage cultures by lipopolysaccharide, muramyl tripeptide, and interleukin-1

The tryptophan decyclizing enzyme indoleamine 2,3-dioxygenase (IDO) was induced in human monocyte-derived macrophages (MDM) treated with human recombinant interferon-β (IFN-β) or interferon-γ (IFN-γ). Treated cells exhibited dose-dependent increases in IDO when assayed 48 hr after treatment. Cells exposed to IFN-γ were observed to exhibit consistently higher peak levels of IDO when compared with cells incubated in the presence of IFN-β. When IFN-β-treated cells were incubated in the presence of specified amounts of bacterial lipopolysaccharide (LPS) or liposome-encapsulated muramyl tripeptide (MTP), peak IDO activity increased such that enzyme activity was comparable to maximal activity observed with IFN-γ-treated cells. LPS and MTP also upregulated IFN-γ-mediated IDO activity when suboptimal amounts of IFN-γ were used. When macrophages were costimulated with various concentrations of human recombinant interleukin 1α (IL-1α), along with either maximum-stimulating amounts of IFN-β or suboptimal amounts of IFN-γ, IDO activity was upregulated in a manner similar to results obtained using the microbial products as stimuli. While neither IL-1α or IL-1β was detected in culture supernatants from macrophages treated with either LPS or MTP (alone or in combination with IFN), IL-1α was detected in cell lysates of macrophages treated with these upregulators. Although neutralizing antibody to IL-1α abolished the upregulatory effect of exogenous IL-1α, it had no effect on upregulation by LPS or MTP. This suggests that although LPS and MTP may induce production of cell-associated IL-1α, upregulation of IDO activity by these agents is independent of IL-1α production and may be mediated through distinct pathways.     

Chaperone SecB: Conformational changes demonstrated by circular dichroism

The chaperone SecB, which is involved in protein export inEscherichia coli, is shown by circular dichroism measurements to contain a high content of-pleated sheets. Prediction of the secondary structure of SecB is in good agreement with the observed content of-sheet. In accordance with the previous studies in which changes in conformation were assessed indirectly [Randall (1992),Science 257, 241–245], here we show that the conformation of SecB changes with the concentration of salt in the milieu and also when SecB interacts with a peptide ligand.Abbreviations ANS 1-anilino-naphthalene-8-sulfonate - CD circular dichroism - NMR nuclear magnetic resonance - CCA convex constraint analysis