Synthesis of polypeptides by microwave heating II. Function of polypeptides synthesized during repeated hydration-dehydration cycles

Summary Polypeptides, synthesized from a mixture of amino acid amides by microwave heating during repeated hydration-dehydration cycles, showed hydrolase- and oxidoreductase-like catalytic activities. Poly(GAVDH), polypeptides synthesized from an equimolar mixture (each 0.1 M) of glycinamide,l-alaninamide,l-valinamide,l-aspartic acid -amide, andl-histidinamide, catalyzed the hydrolysis of PNPAc. The hydrolytic rate of PNPAc with poly(GAVDH) was the quadruple of that ofl-histidine alone. Though the k_cat values of different resulting polypeptides were 10³–10⁶ times less than those of native hydrolases, the K_m value of the polypeptides further containing phenylalanine residues was nearly equal to that of the esterase. This result indicates the presence of hydrophobic interaction between a substrate and the polypeptides. Resulting polypeptides also showed catalytic activity for the reduction of ferricyanide ion [Fe(CN)^3–] with NADH. The polypeptides seemed to have a strong affinity for adenine nucleotides, because the reaction was inhibited by adenine derivatives such as NAD⁺ and AppA. A hypothesis for the emergence of primitive protein enzymes is discussed.     

Influences of root distribution and growth on predicted water uptake and interspecific competition

Summary Root distribution and growth measured in the field were incorporated into a water uptake model for the CAM succulent Agave deserti and its nurse plant Hilaria rigida, a common desert bunchgrass. Agave deserti responds to the infrequent rainfalls of the Sonoran Desert by extending its existing established roots and by producing new roots. Most of such root growth was completed within one month after soil rewetting, total root length of A. deserti increasing by 84% for a seedling and by 58% for a mediumsized plant in the summer. Root growth in the winter with its lower soil temperatures was approximately half as much as in the summer. For a 15-year period, predicted annual root growth of A. deserti varied more than 18-fold because of annual variations in rainfall amount and pattern as well as seasonal variation in soil temperature. Predicted annual water uptake varied 47-fold over the same period. The nurse plant, which is crucial for establishment of A. deserti seedlings, reduced seedling water uptake by 38% during an average rainfall year. Lowering the location of the root system of a medium-sized A. deserti by 0.24 m reduced its simulated annual water uptake by about 25%, reflecting the importance of shallow roots for this desert succulent. Lowering the root system of a medium-sized H. rigida by 0.28 m increased the simulated annual water uptake of an associated A. deserti seedling by 17%, further indicating the influence of root overlap on competition for water.     

Inheritance of antagonistic properties and lytic enzyme activities in sexual crosses of Talaromyces flavus

Two wild-type strains and three benomyl-resistant mutants of the antagonistic ascomycete Talaromyces flavus were crossed in six combinations, two of which yielded hybrid cleistothecia. Parental strains and their ascospore progenies varied in several traits considered to play an important role in the capacity to control soilborne fungal pathogens: extracellular activities of glucose oxidase and cell-wall degrading enzymes, antibiosis towards Verticillium dahliae, and parasitism and biocontrol of Sclerotium rolfsii. A non-Mendelian quantitative mode of inheritance was found for β-1, 3-glucanase and chitinase activities but only the latter exhibited a normal frequency distribution. Some of the progenies exhibited higher glucanase and chitinase activities than those found in the parental strains. Progeny analysis for chitinase, glucanase, cellulase, and glucose oxidase activities revealed no genetic association between any two of these enzymes. Antibiosis was correlated with glucose-oxidase activity in one cross, but not in the other. The ability to reduce bean root rot caused by S. rolfsii was correlated with mycoparasitic activity against S. rolfsii sclerotia in one cross, but not in the other. One out of the 20 progenies tested was able to reduce bean root rot more effectively than its parent strains, thus demonstrating the feasibility of improving a biocontrol agent by conventional breeding.     

植物群落演替与土壤发展之间的关系

本文运用植物群落中植物优势种的重要值与相对应子群落的土壤特征值直接相关联的分析方法,对湖北宜昌大老岭的桦(Betula)、栎(Quercus)、栗(Castanea)七个群落类型的三个演替途径的土壤动态进行分析,结果表明:在植物群落的演替过程中,土壤中的Ca、pH值减少;离子代换量增加;Na、Mg、NH_4+-N、速效P、速效K、有机质的含量是随着物种的变化而呈不同的趋势。因此,植物群落的演替是生态系统的动态过程。     

Antioxidant enzyme activities in embryologic and early larval stages of turbot

The antioxidant enzymes superoxide dismutase (SOD; EC 1.15.1.1), catalase (EC 1.11.1.6), selenium-dependent glutathione peroxidase (SeGPX; EC 1.11.1.9), glutathione reductase (EC 1.6.4.2) and DT-diaphorase (EC 1.6.99.2), plus total GPX activity (sum of SeGPX and Se-independent GPX activities), were studied in 13 500 g supernatants of embryos and 3-day and 11-day post-hatch larvae of turbot Scophthalmus maximus L. SOD activity decreased progressively during development from embryos to 11-day-old larvae, indicative of a decreased need to detoxify superoxide anion radical (O₂⁻). In contrast, catalase, SeGPX and glutathione reductase activities increased progressively from embryos to 11-day-old larvae, indicative of an increased need to metabolize hydrogen peroxide (H₂O₂) and organic peroxides. Consistent with the latter changes, levels of lipid peroxides (i.e. thiobarbituric acid reactive substances) increased 13-fold from embryos to 3-day-old larvae, whilst total peroxidizable lipid was indicated to decrease. Increases were seen for NADPH-dependent DT-diaphorase (after hatching) and total GPX (between 3 and 11 days post-hatch) activities, whilst no change was found in NADH-dependent DT-diaphorase activity. Overall, the results demonstrate a capacity for early life-stages of S. maximus to detoxify reactive oxygen species (O₂⁻ and H₂O₂) and other pro-oxidant compounds (organic peroxides, redox cycling chemicals). Furthermore, qualitative and quantitative antioxidant changes occur during hatching and development, possibly linked to such events as altered respiration rates (SOD changes) and tissue reorganization and development (catalase, SeGPX, lipid peroxidation).     

Changes of the forest-savanna boundary in Brazilian Amazonia during the Holocene revealed by stable isotope ratios of soil organic carbon

The possibility of ecosystem boundary changes in northern Brazilian Amazonia during the Holocene period was investigated using soil organic carbon isotope ratios. Determination of past and present fluctuations of the forest-savanna boundary involved the measurement of natural ¹³C isotope abundance, expressed as ¹³C, in soil organic matter (SOM). SOM ¹³C analyses and radiocarbon dating of charcoal fragments were carried out on samples derived from soil profiles taken along transects perpendicular to the ecotonal boundary. SOM ¹³C values in the upper soil horizons appeared to be in equilibrium with the overlying vegetation types and did not point to a movement of the boundary during the last decades. However, ¹³C values obtained from deeper savanna and forest soil layers indicated that the vegetation type has changed in the past. In current savanna soil profiles, we observed the presence of mid-Holocene charcoals derived from forest species: fire frequency at that time was probably greater, and more extensive savanna may have resulted. Isotope data and the presence of these charcoals thus suggest that the forest-savanna boundary has shifted significantly in the recent Holocene period, forest being more extensive during the early Holocene than today. During the middle Holocene, the forest could have strongly regressed, and fires appeared, with a maximum development of the savanna vegetation. At the beginning of the late Holocene, the forest may have invaded a part of this savanna, and fires occurred again.     

Causal connection between detoxification enzyme activity and consumption of a toxic plant compound

Insect herbivores can increase their detoxification activities against a particular plant poison in response to prolonged ingestion of the same compound. For example, larval tobacco hornworms (Manduca sexta) experience a dramatic increase in cytochrome P450 activity against nicotine after ingesting nicotine. While it is generally assumed that this induction process permits increased consumption of toxic plant tissues, we are not aware of any direct experimental support for this assumption. Using a two-tiered approach, we examined the functional significance of P450 induction to M. sexta larvae ingesting a toxic but non-deterrent concentration of nicotine. First, we related the time-course of P450 induction in midgut microsomes to changes in nicotine consumption. When offered a nicotine diet, larvae failed to show a significant increase in consumption before 36 h, which was coincident with the time-course of the induction of midgut P450 activities against aldrin and nicotine. Second, we determined whether inhibiting the induced P450 activities affected nicotine consumption. We found that the increase in nicotine consumption following the induction of nicotine metabolism could be strongly inhibited by treatment with piperonyl butoxide, which by itself did not inhibit consumption. These results provide direct evidence for a causal connection between P450-mediated detoxification activity and consumption of a toxic plant compound.Abbreviation PB piperonyl butoxide     

The other kind of biological NMR—Studies of enzyme-substrate interactions

NMR spectroscopy has proved to be a valuable tool in the study of the interactions between enzymes and their substrates. The kinds of structural and dynamic information which can be obtained are illustrated by studies of three enzymes involved in drug metabolism. Cytochromes P₄₅₀ play a crucial role in metabolism of a wide range of exogenous chemicals. NMR has been used to measure distances from the haem iron of the cytochrome to protons of the bound substrate, leading to detailed structural models for the enzyme-substrate complexes. The other two enzymes, chloramphenicol acetyltransferase and β-lactamase, are responsible for bacterial resistance to specific antibiotics. In chloramphenicol acetyltransferase, NMR has been used to determine the conformation of coenzyme A bound to the enzyme, while in the case of β-lactamase the pK of a specific lysine residue at the active site has been determined, providing valuable information on the catalytic mechanism. Special issue dedicated to Dr. Herman Bachelard.     

Temporal and ontogenetic aspects of protein induction in foliage of the tomato, Lycopersicon esculentum

Damage to foliage of the tomato, Lycopersicon esculentum, causes the induction of proteinase inhibitors and of the oxidative enzymes polyphenol oxidase, peroxidase, and lipoxygenase. The time courses of induction of these proteins by feeding of two caterpillar species (Manduca sexta and Helicoverpa zea) were studied in a series of experiments. In another series of experiments, the effects of plant age on the inducibility of these proteins were studied. In the time course experiments, induction of proteinase inhibitors and oxidative enzymes in the damaged leaflet was rapid, with higher protein activities evident in damaged leaflets within 12–24 h of damage, depending on the enzyme and the species of insect used to damage the plant. Systemic induction of proteinase inhibitors was also rapid, but systemic induction of polyphenol oxidase was delayed relative to systemic induction of proteinase inhibitors, possibly because high constitutive polyphenol oxidase activities obscured expression of systemic induction at earlier time points. Lipoxygenase and peroxidase were not induced systemically. Induction of all proteins persisted for at least 21 days. In the phenology experiments, inducibility of all proteins decreased in magnitude and was less consistent as plants aged. The results of these experiments exemplify the numerous constraints on induction in tomato plants. Knowledge of these physiological constraints is important to an understanding of the ecological role and causal basis of induced resistance.     

Distribution and activity of microsomal NADPH-dependent monooxygenases and amino acid decarboxylases in cruciferous and non-cruciferous plants, and their relationship to foliar glucosinolate content

The NADPH-dependent conversion of amino acids to their aldoximes is an initial step in glucosinolate biosynthesis. A number of microsomal aldoxime-forming monooxygenase activities were detected in leaves from a variety of glucosi-nolate-containing species, whereas barley, bean and tobacco leaves did not contain any such activities. The substrates for these monooxygenases in each species largely correlated with the spectrum of glucosinolates found in that species. No activity was detected that metabolized homomethionine (supposed precursor of 2-propenylglucosinolate [sinigrin]), even in species where sinigrin was the major glucosinolate. In Sinapis species containing hydroxybenzylglucosinolate (sinalbin), activity with L-Tyr was detected, whereas Brassica species containing sinalbin had no such activity. However, these Brassicas did contain an L-Phe monooxygenase activity. Partial characterization of the monooxygenases indicated that in Brassica species, Nasturtium officinalis and Raphanus sativus these resembled the flavin-linked monooxygenases previously found in oilseed rape (Brassica napus) and Chinese cabbage (Brassica campestris). The L-Tyr-dependent activity in Sinapis species, and the L-Phe-dependent activity in Tropacolum majus, had characteristics of cytochrome P450-type enzymes. No similarity was found with any other known amino acid metabolizing enzymes (including decarboxylases, amino acid oxidases and diamine/polyamine oxidases).