Greenhouse warming by nitrous oxide and methane in the Proterozoic Eon

An anoxic, sulfidic ocean that may have existed during the Proterozoic Eon (0.54-2.4 Ga) would have had limited trace metal abundances because of the low solubility of metal sulfides. The lack of copper, in particular, could have had a significant impact on marine denitrification. Copper is needed for the enzyme that controls the final step of denitrification, from N(2) O to N(2) . Today, only about 5-6% of denitrification results in release of N(2) O. If all denitrification stopped at N(2) O during the Proterozoic, the N(2) O flux could have been 15-20 times higher than today, producing N(2) O concentrations of several ppmv, but only if O(2) levels were relatively high (>0.1 PAL). At lower O(2) levels, N(2) O is rapidly photodissociated. Methane concentrations may also have been elevated during this time, as has been previously suggested. A lack of dissolved O(2) and sulfate in the deep ocean could have produced a high methane flux from marine sediments, as much as 10-20 times today's methane flux from land. The photochemical lifetime of CH(4) increases as more CH(4) is added to the atmosphere, so CH(4) concentrations of up to 100 ppmv are possible during this time. The combined greenhouse effect of CH(4) and N(2) O could have provided up to 10° of warming, thereby keeping the surface warm during the Proterozoic without necessitating high CO(2) levels. A second oxygenation event near the end of the Proterozoic would have resulted in a reduction in both atmospheric N(2) O and CH(4) , perhaps triggering the Neoproterozoic "Snowball Earth" glaciations.     

Evolutionary timing of the origins of mesophilic sulphate reduction and oxygenic photosynthesis: a phylogenomic dating approach

Until recently, the deep‐branching relationships in the bacterial domain have been unresolved. A new phylogenetic approach (termed compartmentalization) was able to resolve these deep‐branching relationships successfully by using a large number of genes from whole genome sequences and by reducing long branch attraction artefacts. This new, well‐resolved phylogenetic tree reveals the evolutionary relationships between diverse bacterial groups that leave important traces in the geological record. It shows that mesophilic sulphate reducers originated before the Cyanobacteria, followed by the origination of sulphur‐ and pyrite‐oxidizing bacteria after oxygen became available in the biosphere. This evolutionary pattern mirrors a similar pattern in the Palaeoproterozoic geological record. Sulphur isotopic fractionation records indicate that large‐scale bacterial sulphate reduction began in marine environments around 2.45 billion years ago (Ga), followed by rapid oxygenation of the atmosphere about 2.3 or 2.2 Ga. Oxygenation was then followed by increasing oceanic sulphate concentrations (probably owing to pyrite oxidation and continental weathering), which then resulted in the disappearance of banded iron formations by 1.8 Ga. The similarity between the phylogenetic and geological records suggests that the geochemical changes observed on the Palaeoproterozoic Earth were caused by major origination events in the mesophilic bacteria, and that these geochemical changes then caused additional origination events, such as aerobic respiration. If so, then constraints on divergence dates can be established for many microbial groups, including the Cyanobacteria, mesophilic bacteria, mesophilic sulphate reducers, methanotrophs, several anoxygenic phototrophs, as well as for mitochondrial endosymbiosis. These dates may also help to explain a large number of other changes in the geological record of the Neoarchean and Palaeoproterozoic Earth. This hypothesis, however, does not agree with the finding of cyanobacterial and eukaryote lipids at 2.7 Ga, and suggests that further work needs to be done to elucidate the discrepancies in both these areas.     

Atmospheric composition and climate on the early Earth

Oxygen isotope data from ancient sedimentary rocks appear to suggest that the early Earth was significantly warmer than today, with estimates of surface temperatures between 45 and 85 degrees C. We argue, following others, that this interpretation is incorrect-the same data can be explained via a change in isotopic composition of seawater with time. These changes in the isotopic composition could result from an increase in mean depth of the mid-ocean ridges caused by a decrease in geothermal heat flow with time. All this implies that the early Earth was warm, not hot.A more temperate early Earth is also easier to reconcile with the long-term glacial record. However, what triggered these early glaciations is still under debate. The Paleoproterozoic glaciations at approximately 2.4Ga were probably caused by the rise of atmospheric O2 and a concomitant decrease in greenhouse warming by CH4. Glaciation might have occurred in the Mid-Archaean as well, at approximately 2.9Ga, perhaps as a consequence of anti-greenhouse cooling by hydrocarbon haze. Both glaciations are linked to decreases in the magnitude of mass-independent sulphur isotope fractionation in ancient rocks. Studying both the oxygen and sulphur isotopic records has thus proved useful in probing the composition of the early atmosphere.     

Geological and trace element evidence for a marine sedimentary environment of deposition and biogenicity of 3.45 Ga stromatolitic carbonates in the Pilbara Craton, and support for a reducing Archaean ocean

Bedded carbonate rocks from the 3.45 Ga Warrawoona Group, Pilbara Craton, contain structures that have been regarded either as the oldest known stromatolites or as abiotic hydrothermal deposits. We present new field and petrological observations and high‐precision REE + Y data from the carbonates in order to test the origin of the deposits. Trace element geochemistry from a number of laminated stromatolitic dolomite samples of the c. 3.40 Ga Strelley Pool Chert conclusively shows that they precipitated from anoxic seawater, probably in a very shallow environment consistent with previous sedimentological observations. Edge‐wise conglomerates in troughs between stromatolites and widespread cross‐stratification provide additional evidence of stromatolite construction, at least partly, from layers of particulate sediment, rather than solely from rigid crusts. Accumulation of particulate sediment on steep stromatolite sides in a high‐energy environment suggests organic binding of the surface. Relative and absolute REE + Y contents are exactly comparable with Late Archaean microbial carbonates of widely agreed biological origin. Ankerite from a unit of bedded ankerite–chert couplets from near the top of the stratigraphically older (3.49 Ga) Dresser Formation, which immediately underlies wrinkly stromatolites with small, broad, low‐amplitude domes, also precipitated from anoxic seawater. The REE + Y data of carbonates from the Strelley Pool Chert and Dresser Formation contrast strongly with those from siderite layers in a jasper–siderite–Fe‐chlorite banded iron‐formation from the base of the Panorama Formation (3.45 Ga), which is clearly hydrothermal in origin. The geochemical results, together with sedimentological data, strongly support: (1) deposition of Dresser Formation and Strelley Pool Chert carbonates from Archaean seawater, in part as particulate carbonate sediment; (2) biogenicity of the stromatolitic carbonates; (3) a reducing Archaean atmosphere; (4) ongoing extensive terrestrial erosion prior to ～3.45 Ga.     

Quinone toxicity in hepatocytes: studies on mitochondrial Ca2+ release induced by benzoquinone derivatives

Hepatocyte cytotoxicity caused by substituted benzoquinones was associated with increased cytosolic Ca2+ concentration. p-Benzoquinone-induced hepatotoxicity was enhanced when the hepatocytes were loaded with Ca2+ by preincubation with ATP. A similar order of potency of the substituted benzoquinones in releasing Ca2+ from isolated mitochondria and inducing hepatocyte cytotoxicity was found; in decreasing order, this was 2-Br-, unsubstituted-, 2-CH3-, 2,6-(CH3O)2-, 2,6-(CH3)2-, 2,5-(CH3)2-, 2,3,5-(CH3)3-, and 2,3,5,6-(CH3)4-benzoquinones (duroquinone). The cellular products of quinone metabolism, hydroquinones and glutathione conjugates, did not cause mitochondrial Ca2+ release. Benzoquinone-induced mitochondrial Ca2+ release was preceded by GSH conjugate formation and NAD(P)H oxidation but followed by mitochondrial swelling. With duroquinone, a slow GSH and NADPH oxidation preceded Ca2+ release, but GSH oxidation did not occur with Se-deficient mitochondria lacking glutathione peroxidase activity. Cyanide-insensitive respiration was also observed with duroquinone but not with benzoquinone, suggesting that duroquinone undergoes redox cycling. GSH was depleted by both arylation and oxidation with 2,6-(CH3O)2-, 2,6-(CH3)2-, 2,5(CH3)2-, and 2,3,5-(CH3)3-benzoquinones. Benzoquinone concentrations that totally depleted GSH did not cause Ca2+ release until intramitochondrial NAD(P)H was oxidized. Ca2+ release was also prevented when NAD(P)H generation was stimulated by the presence of isocitrate or 3-hydroxybutyrate. This suggests that mitochondrial Ca2+ release is associated with NAD(P)H oxidation catalyzed by NADH dehydrogenase with benzoquinone or by the glutathione peroxidase-glutathione reductase system with duroquinone.     

Seaweeds in cold seas: evolution and carbon acquisition

Much evidence suggests that life originated in hydrothermal habitats, and for much of the time since the origin of cyanobacteria (at least 2.5 Ga ago) and of eukaryotic algae (at least 2.1 Ga ago) the average sea surface and land surface temperatures were higher than they are today. However, there have been at least four significant glacial episodes prior to the Pleistocene glaciations. Two of these (approx. 2.1 and 0.7 Ga ago) may have involved a 'Snowball Earth' with a very great impact on the algae (sensu lato) of the time (cyanobacteria, Chlorophyta and Rhodophyta) and especially those that were adapted to warm habitats. By contrast, it is possible that heterokont, dinophyte and haptophyte phototrophs only evolved after the Carboniferous-Permian ice age (approx. 250 Ma ago) and so did not encounter low (相似文献   

Forest Management Impacts on Greenhouse Gas Fluxes from Riparian Soils Along Headwater Streams

Ecosystems - Increasing concentrations of atmospheric greenhouse gases (GHGs; CO2, CH4, N2O) cause climate change. Depending on the conditions, soils have the potential to store carbon or to be a...     

Relay cropping for improved air and water quality

Using plants to extract excess nitrate from soil is important in protecting against eutrophication of standing water, hypoxic conditions in lakes and oceans, or elevated nitrate concentrations in domestic water supplies Global climate change issues have raised new concerns about nitrogen (N) management as it relates to crop production even though there may not be an immediate threat to water quality. Carbon dioxide (CO2) emissions are frequently considered the primary cause of global climate change, but under anaerobic conditions, animals can contribute by expelling methane (CH4) as do soil microbes. In terms of the potential for global climate change, CH4 is approximately 25 times more harmful than CO2. This differential effect is minuscule compared to when nitrous oxide (N2O) is released into the atmosphere because it is approximately 300 times more harmful than CO2. N2O losses from soil have been positively correlated with residual N (nitrate, NO3-) concentrations in soil. It stands to reason that phytoremediation via nitrate scavenger crops is one approach to help protect air quality, as well as soil and water quality. Winter wheat was inserted into a seed corn/soybean rotation to utilize soil nitrate and thereby reduce the potential for nitrate leaching and N2O emissions. The net effect of the 2001-2003 relay cropping sequence was to produce three crops in two years, scavenge 130 kg N/ha from the root zone, produce an extra 2 Mg residue/ha, and increase producer profitability by approximately 250 dollars/ha.     

Methylammonium transport in Anacystis nidulans R-2

Methylammonium was taken up rapidly by illuminated cells of Anacystis nidulans R-2, leading to internal concentrations of 1.3 +/- 0.1 mM within 1 min, and a gradient of up to 200 between the cells and medium. Accumulation of 14CH3NH3+ required at least 5 mM NaCl, but the uptake rate was independent of medium pH between 6.5 and 9. The kinetics of uptake could be resolved into an initial fast phase lasting less than 1 min (approximate Km, 7.2 microM; Vmax, 12.5 nmol min-1 mg of protein-1 at 15 degrees C). A second, slower phase associated with product formation was eliminated by preincubation with methionine sulfoximine, a specific inhibitor of glutamine synthetase; the rapid phase was unaffected by this treatment. Ammonium ions competed with 14CH3NH3+ for entry, and addition of 5 microM NH4+ or 100 microM CH3NH3+ released 14CH3NH3+ accumulated during the rapid phase of entry. Small additions of NH4+ made at the same time as additions of 14CH3NH3+ delayed the start of radioactivity uptake by a time which corresponded accurately with the period needed for the complete removal of the added NH4+. The effects of inhibitors on accumulation and carbocyanine dye fluorescence suggest that ATP-dependent membrane potential was needed to drive 14CH3NH3+ transport. Spheroplasts were as active as whole cells in accumulating NH4+ and 14CH3NH3+, indicating that soluble periplasmic components are not involved in the translocation. Some significant differences between the translocation of 14CH3NH3 and that of NH4+ were observed: growth with NH4+ in place of NO3- repressed 14CH3NH3+ accumulation ability without affecting the NH4+ uptake rate Na+ was not required for NH4+ uptake, and concentration of KCl inhibitory with 14C3NH3+ did not reduce NH4+ uptake.     

Endogenous methanogenesis stimulates oxidation of atmospheric CH(4) in alpine tundra soil

Experiments were done to test the hypothesis that atmospheric CH(4) oxidizers in a well-drained alpine tundra soil are supported by CH(4) production from anaerobic microsites in the soil. Soil was subjected to 22 days of anaerobic conditions with elevated H(2) and CO(2) in order to stimulate methanogenesis. This treatment stimulated subsequent atmospheric CH(4) consumption, probably by increasing soil methanogenesis. After removal from anaerobic conditions, soils emitted CH(4) for up to 6 h, then oxidized atmospheric CH(4) at 111 (+/- 5.7) pmol (g dry weight)(-1) h(-1), which was more than 3 times the rate of control soils. Further supporting our hypothesis, additions of lumazine, a highly specific inhibitor of methanogenesis, prevented the stimulation of atmospheric CH(4) oxidation by the anaerobic treatment. The method used to create anaerobic conditions with elevated H(2) and CO(2) also elevated headspace CH(4) concentrations. However, elevated CH(4) concentrations under aerobic conditions did not stimulate CH(4) oxidation as much as preexposure to H(2) and CO(2) under anaerobic conditions. Anaerobic conditions created by N(2) flushing did not stimulate atmospheric CH4 oxidation, probably because N2 flushing inhibited methanogenesis by removing necessary precursors for methane production. We conclude that anaerobic conditions with elevated H(2) and CO(2) stimulate atmospheric CH(4) oxidation in this dry alpine tundra soil by increasing endogenous CH(4) production. This effect was prevented by inhibiting methanogenesis, indicating the importance of endogenous CH(4) production in a CH(4-) consuming soil.     

Simultaneous methanogenesis and oxygen reduction by Methanobrevibacter cuticularis at low oxygen fluxes

Methanogenic Archaea are often encountered in habitats that are not entirely anoxic in space or time. Recent biochemical and genomic studies have revealed the capacity of methanogens to reduce molecular oxygen. O(2) reduction by Methanobrevibacter species was investigated. Cell suspensions incubated in agar tubes under a headspace of H(2)-CO(2) and increasing concentrations of O(2) formed a distinct growth band, which coincided with the oxic-anoxic interface and indicated that the influx of O(2) into the band was balanced by its consumption. However, in batch cultures methanogenesis ceased as soon as traces of O(2) were added. Focusing on Methanobrevibacter cuticularis, a species colonizing the microoxic gut epithelium of termites, a diffusion-limited setup was used that allowed the exposure of dense cell suspensions to controlled O(2) fluxes. Here, Methanobrevibacter cuticularis was capable of simultaneous CH(4) production and O(2) consumption. Low O(2) fluxes (10% of the CH(4) production rate) had virtually no influence on methanogenesis [4.5 micromol CH(4) (mg dry wt)(-1) h(-1)], whereas higher O(2) fluxes (up to 30% of the initial CH(4) production rate) caused a reversible decrease in methanogenesis, which was accompanied by a reversible, partial conversion of coenzyme F(420) to factor F(390). The maximum O(2) reduction rate [4.8 micromol O(2) (mg dry wt)(-1) h(-1)] that could be maintained over extended time periods (>30 min) was similar to the CH(4) production rate under anoxic conditions.     

Differential Inhibition by Allylsulfide of Nitrification and Methane Oxidation in Freshwater Sediment

Addition of nitrapyrin, allylthiourea, C(inf2)H(inf2), and CH(inf3)F to freshwater sediment slurries inhibited CH(inf4) oxidation and nitrification to similar extents. Dicyandiamide and allylsulfide were less inhibitory for CH(inf4) oxidation than for nitrification. Allylsulfide was the most potent inhibitor of nitrification, and the estimated 50% inhibitory concentrations for this process and CH(inf4) oxidation were 0.2 and 121 (mu)M, respectively. At a concentration of 2 (mu)M allylsulfide, growth and CH(inf4) oxidation activity of Methylosinus trichosporium OB3b were not inhibited. Allylsulfide at 200 (mu)M inhibited the growth of M. trichosporium by approximately 50% but did not inhibit CH(inf4) oxidation activity. Nitrite production by cells of M. trichosporium was not significantly affected by allylsulfide, except at a concentration of 2 mM, when growth and CH(inf4) oxidation were also inhibited by about 50%. Methane monooxygenase activity present in soluble fractions of M. trichosporium was not inhibited significantly by allylsulfide at either 200 (mu)M or 2 mM. These results suggest that the partial inhibition of CH(inf4) oxidation in sediment slurries by high allylsulfide concentrations may be caused by an inhibition of the growth of methanotrophs rather than an inhibition of methane monooxygenase activity specifically. We conclude that allylsulfide is a promising tool for the study of interactions of methanotrophs and nitrifiers in N cycling and CH(inf4) turnover in natural systems.     

中亚热带米槠天然林土壤甲烷吸收速率季节变化

以福建省建瓯市万木林自然保护区米槠天然林为对象,定位观测了土壤甲烷吸收速率(V_CH4)的季节变化.结果表明：米槠天然林土壤V_CH4的季节变化表现出夏秋季高于冬春季的趋势,最大值(95.13 μg·m^-2·h^-1)出现在初秋(9月),最小值(9.13 μg·mμg·m^-2·h^-1)出现在初春(3月).土壤全年均为甲烷汇.随土壤温度和含水量的增加, V_CH4分别呈增加和降低趋势,但V_CH4与土壤温度和土壤含水量的相关性均不显著.米槠天然林土壤甲烷年通量为3.93 kg·hm^-2·a^-1,高于全球天然林土壤甲烷年通量的平均水平(2.4 kg·hm^-2·a^-1)和亚洲地区热带天然林土壤甲烷年通量(2.07 kg·hm^-2·a^-1),低于亚洲地区温带天然林的土壤甲烷年通量(8.12 kg·hm^-2·a^-1).     

华北高于庄组硅化微体化石组合的古环境

华北中元古代高于庄组 ( 1 4- 1 5亿年 )的硅质叠层石中保存了完好的多种微生物化石。从这些叠层石的微细构造分析 ,成岩早期硅质矿物的交代作用为微体化石的原位保存起了很重要的作用。除部分居住者和浮游的分子外 ,两种丝状蓝藻 ( Siphonophycus inornatum和Eoschizothric composita)和两种球状蓝藻 ( Coccostratusdispergens和 Eoentophysalisbelcherensis)是这些藻席的主要建造者。以球状蓝藻 Eoentophysalis为主的藻席可能发育于潮下高能环境中 ;而以多种丝状蓝藻为主的藻席可能反映了当时的沉积环境为中—高潮间带的局部静止小水体     

Auslösung der Blütenbildung durch Cycloheximid im Kurztag bei der Langtagpflanze Hyoscyamus niger

Summary Cycloheximide (CH) was applied selectively either to the shoot apex or by infiltration to the leaves of the long-day plant Hyoscyamus niger in order to investigate whether this inhibitor has an effect on the synthesis of a floral stimulus in the leaves. Treatment of the shoot apex with CH caused inhibition of the photoperiodic induction. In contrast, when CH was applied to leaves, initiation of flowering was observed under short-day conditions. The drug yielded optimum initiating effects at concentrations of 10^-5-3·10^-5 M, inducing flowering of almost 60% of the plants. Daily infiltration over a period of up to 4 days decreased the rate of flower initiation. The effect of CH was shown to be additive to a photoperiodic induction, even to a sub-threshold induction, but not to 2-thiouracil mediated induction. In no case did the presence of additional untreated leaves on the plants suppress CH-mediated flower induction. Treatment of the leaves with chloramphenicol (10^-6-2^-10^-4 M) or puromycin (5·10^-6-2·10^-4 M) caused no initiating response. The results are interpreted to mean that the presence of CH in the leaves may lead to the synthesis of a floral stimulus also under short-day conditions. This finding is similar to that reported previously in the case of the inductive effect of 2-thiouracil.

---

Folgende Abkürzungen wurden verwendet 2-TU 2-Thiouracil - CH Cycloheximid - LT Langtag - DL Dauerlicht Herrn Prof. Dr. L. Brauner in Verehrung und Dankbarkeit zum 75. Geburtstag gewidmet.     

Coupling of methyl coenzyme M reduction with carbon dioxide activation in extracts of Methanobacterium thermoautotrophicum

The stimulation of carbon dioxide reduction to methane by addition of 2-(methylthio)ethanesulfonate (CH3-S-CoM) to cell extracts of Methanobacterium thermoautotrophicum was investigated. Similar stimulation of CO2 reduction by CH3-S-CoM was found for cell extracts of Methanobacterium bryantii and Methanospirillum hungatei. The CH3-S-CoM requirement could be met by the methanogenic precursors formaldehyde, serine, or pyruvate, or by 2-(ethylthio)ethanesulfonate (CH3CH2-S-CoM), but not by other coenzyme M derivatives. Efficient reduction of CO2 to CH4 was favored by low concentrations of CH3-S-CoM and high concentrations of CO2. Sulfhydryl compounds were identified as effective inhibitors of CO2 reduction. Both an allosteric model and a free-radical model for the mechanism of CO2 activation and reduction are discussed.     

Fermentation pattern of methanogenic degradation of rice straw in anoxic paddy soil

The anaerobic degradation of different fractions of rice straw in anoxic paddy soil was investigated. Rice straw was divided up into stem, leaf sheath and leaf blade. The different straw fractions were mixed with paddy soil and incubated under anoxic conditions. Fermentation of straw components started immediately and resulted in transient accumulation of acetate, propionate, butyrate, isobutyrate, valerate, isovalerate and caproate with much higher concentrations in the presence than in the absence of straw. Also some unidentified compounds with UV absorption could be detected. The maximum concentrations of these compounds were different when using different straw fractions, suggesting differences in the degradation pathway of these straw fractions during the early phase of incubation, i.e. with Fe(III) and sulfate serving as oxidants. When concentrations of the intermediates decreased to background values, CH(4) production started. Rates of CH(4)unamended soil. During the methanogenic phase, the percentage contribution of fermentation products to CH(4) production was determined by inhibition with 2-bromoethanesulfonate (BES). Acetate (48-83%) and propionate (18-28%) were found to be the main intermediates of the carbon flow to CH(4), irrespective of the fraction of the rice straw or its absence. Mass balance calculations showed that 84-89% of CH(4) was formed via acetate in the various incubations. Radiotracer experiments showed that 11-27% of CH(4) was formed from H(2)/CO(2), thus confirming that acetate contributed 73-89% to methanogenesis. Our results show that the addition of rice straw and the fraction of the straw affected the fermentation pattern only in the early phase of degradation, but had no effect on the degradation pathway during the later methanogenic phase.     

Chronic hypoxia-induced spontaneous and rhythmic contractions in the rat main pulmonary artery

The effect of chronic hypoxia (CH; 1-4 wk) on the electromechanical properties of the rat main pulmonary artery (MPA) was investigated. MPA rings obtained from rats exposed for 14 days to hypobaric (50.5 kPa) CH exhibited spontaneous and rhythmic contractions (SRCs) that were never observed in control (normoxic) rats. SRCs were unaffected by tetrodotoxin, phentolamine, BQ-123 and BQ-788, N-nitro-L-arginine methyl ester, or endothelium removal. CH depolarized smooth muscle cells from -58.8 +/- 9 to -38.6 +/- 5.4 mV and increased the resting cytosolic Ca2+ concentration from 67.3 +/- 11.9 to 112.5 +/- 16.4 nM. CH also induced spontaneous spikelike depolarizations. All of these effects were inhibited by external Ca2+ removal or nifedipine (1 microM). Moreover, depletion of intracellular Ca2+ stores with ryanodine (1-5 microM) or cyclopiazonic acid (3 microM) progressively attenuated SRCs. This study demonstrates that CH switches the MPA from a quiescent to a spontaneously active mechanical state. Finally, the fact that SRCs precede the development of right ventricle hypertrophy and disappear when this hypertrophy reaches a maximal value (after 3-4 wk of CH) suggests that SRCs may play a role in the adaptive process of the pulmonary circulation to CH.     

Crystal structures of alpha-mercaptoacyldipeptides in the thermolysin active site: structural parameters for a Zn monodentation or bidentation in metalloendopeptidases.

Three alpha-mercaptoacyldipeptides differing essentially in the size of their C-terminal residues have been crystallized in the thermolysin active site. A new mode of binding was observed for 3 [HS-CH(CH(2)Ph)CO-Phe-Tyr] and 4 [HS-CH((CH(2))(4)CH(3))CO-Phe-Ala], in which the mercaptoacyl moieties act as bidentates with Zn-S and Zn-O distances of 2.3 and 2.4 A, respectively, the side chains fitting the S(1), S(1)', and S(2)' pockets. Moreover, a distance of 3.1 A between the sulfur atom and the OE1 of Glu(143) suggests that they are H-bonded and that one of these atoms is protonated. This H-bond network involving Glu(143), the mercaptoacyl group of the inhibitor, and the Zn ion could be considered a "modified" transition state mimic of the peptide bond hydrolysis. Due to the presence of the hindering (5-phenyl)proline, the inhibitor HS-CH(CH(2)Ph)CO-Gly-(5-Ph)Pro (2) interacts through the usual Zn monodentation via the thiol group and occupancy of S(1)' and S(2)' subsites by the aromatic moieties, the proline ring being outside the active site. The inhibitory potencies are consistent with these structural data, with higher affinities for 3 (4.2 x 10(-)(8) M) and 4 (4.8 x 10(-)(8) M) than for 2 (1.2 x 10(-)(6) M). The extension of the results, obtained with thermolysin being considered as the model of physiological zinc metallopeptidases, allows inhibitor-recognition modes for other peptidases, such as angiotensin converting enzyme and neutral endopeptidase, to be proposed and opens interesting possibilities for the design of new classes of inhibitors.     

A study on the reactions of tubercidin triphosphate with myosin and actomyosin.

The triphosphate ester of tubercidin (tubercidin triphosphate, TuTP) was synthesized. This is an analog of ATP in which a CH group replaces the N-7 of the adenine ring. The rate of TuTP hydrolysis by myosin in the presence of Mg2+ was the same as that of ATP in the 10(-7)-10(-3) M range, whereas the increment in the optical density of myosin ihe 290mmu region caused by TuTP was twice that caused by ATP. TuTP is hydrolyzed by actomyosin faster than ATP, the value of Vmax being about 4 times larger while the Km values were of the same order of magnitude. The rate of superprecipitation induced by TuTP was 50% of that caused by ATP at nucleotide concentrations of 3-60 muM. A similar difference was observed with respect to the rate of tension development by glycerol-extracted rabbit psoas fibers upon addition of these two substances. Substitution of ADP by tubercidin diphosphate (TuDP) in F-actin did not affect the rate of superprecipitation or enzymic activity of actomyosin.