Ratio Of Enzyme Activity Research Articles

Three-Enzyme Phosphorylase Cascade for Integrated Production of Short-Chain Cellodextrins.

Cellodextrins are linear β-1,4-gluco-oligosaccharides that are soluble in water up to a degree of polymerization (DP) of ≈6. Soluble cellodextrins have promising applications as nutritional ingredients. A DP-controlled, bottom-up synthesis from expedient substrates is desired for their bulk production. Here, a three-enzyme glycoside phosphorylase cascade is developed for the conversion of sucrose and glucose into short-chain (soluble) cellodextrins (DP range 3-6). The cascade reaction involves iterative β-1,4-glucosylation of glucose from α-glucose 1-phosphate (αGlc1-P) donor that is formed in situ from sucrose and phosphate. With final concentration and yield of the soluble cellodextrins set as targets for biocatalytic synthesis, three major factors of reaction efficiency are identified and partly optimized: the ratio of enzyme activity, the ratio of sucrose and glucose, and the phosphate concentration used. The efficient use of the phosphate/αGlc1-P shuttle for cellodextrin production is demonstrated and the soluble product at 40g L-1 is obtained under near-complete utilization of the donor substrate offered (88mol% from 200mm sucrose). The productivity is 16g(L h)-1 . Through a simple two-step route, the soluble cellodextrins are recovered from the reaction mixture in ≥95% purity and ≈92% yield. Overall, this study provides the basis for their integrated production.

Characteristics of soil enzyme stoichiometry along an altitude gradient on Qinghai-Tibet Pla-teau alpine meadow, China

Soil enzymes play critical roles in material cycle and energy flow of ecosystems. Understanding soil enzyme activities is of great significance for exploring ecosystem functions. In this study, we investigated soil enzyme activities, stoichiometry and their driving factors at six different altitudes (4300-5100 m) on Qinghai-Tibet Plateau alpine meadow using Biolog microplate analysis. The results showed that β-1,4-glucosidase (βG) closely related to C cycle, β-1,4-N-acetylglucosaminidase (NAG) and L-leucine aminopeptidase (LAP) closely related to N cycle and the activity of acid phosphatase (AP), which was closely related to P cycle, all exhibited unimodal trends with increasing altitude, with the order of 4800 m＞4950 m＞4400 m＞4650 m＞5100 m＞4300 m. Soil N:P enzyme activity ratio showed the same trend as soil enzyme activity, and reached the highest value at 4950 m, however, soil C:N and C:P enzyme activities ratios increased along the altitude. Pearson correlation analysis showed that SOC, TN and soil water content were significantly positively correlated with the activities of four types of enzymes. Mean annual precipitation was significantly negatively associated with the activities of NAG and AP. Mean annual precipitation, mean annual temperature, Shannon diversity, vegetation richness, vegetation coverage and TN affected ratios of soil C:P and N:P enzymes. Soil C:N activity ratio correlated with mean annual temperature, mean annual precipitation, vegetation richness, vegetation coverage, SOC and TN. In summary, soil enzyme activities and stoichiometry had remarkable difference along the altitude gradient on Qinghai-Tibet Plateau alpine meadow, with certain N limitation in high altitude areas. Soil water content, TN, SOC, mean annual precipitation and mean annual temperature were key factors driving such differences.

Устойчивость к негативным воздействиям и соотношение активности ферментов энергетического обмена в тканях черноморских моллюсков Mytilus galloprovincialis Lamarck, 1819 и Anadara kagoshimensis (Tokunaga, 1906)

Определение соотношения активности ферментов энергетического обмена малатдегидрогеназы (МДГ, 1.1.1.37) и лактатдегидрогеназы (ЛДГ, 1.1.1.27) позволяет получить интегральную оценку физиологического состояния объекта исследования в ответ на воздействия разной природы. Цель работы ― сравнить изменение величины отношения МДГ/ЛДГ в тканях двустворчатых моллюсков: аборигенной Mytilus galloprovincialis Lamarck, 1819 и успешного вселенца Anadara kagoshimensis (Tokunaga, 1906), в лабораторных условиях под влиянием гипоксии, аноксии, токсиканта полихлорбифенила, сероводородного заражения и длительного содержания в аквариуме без кормления. Половозрелые моллюски собраны в районе г. Севастополя. Длина раковины мидии составляла 45–62 мм, анадары ― 27–49 мм. Активность ферментов измеряли спектрофотометрически (при 340 нм и 25 °C) по скорости окисления НАДН в цитоплазме тканей (мышцы, гепатопанкреас, жабры). Как правило, под воздействием негативных факторов активность ЛДГ снижалась значительно (на 36–80 %), активность МДГ оставалась стабильной, а коэффициент МДГ/ЛДГ в тканях обоих видов моллюсков увеличивался в 1,5–4 раза. Отмечено, что у гемоглобинсодержащей анадары коэффициент МДГ/ЛДГ был на порядок ниже, чем у мидии, как в контроле, так и в опыте. Сопоставление с литературными данными показало, что величина отношения МДГ/ЛДГ устойчивой к гипоксии анадары такая же низкая, как у оксифильных гидробионтов, в частности гребешка Mizuhopecten yessoensis (Jay, 1857) и ракообразных Crangon crangon (Linnaeus, 1758) и Carcinus aestuarii Nardo, 1847. По-видимому, низкое значение коэффициента МДГ/ЛДГ отражает способность анадары сохранять высокий уровень окислительных процессов в тканях благодаря значительному пулу соединений (эритроцитарный гемоглобин, каротиноиды, глутатион), поддерживающих аэробный процесс и осуществляющих антиоксидантную защиту. Величина коэффициента МДГ/ЛДГ может быть использована для оценки степени оксигенации тканей моллюсков в норме и в условиях гипоксии различного происхождения при мониторинговых исследованиях.

Phosphorus addition accelerates fine root decomposition by stimulating extracellular enzyme activity in a subtropical natural evergreen broad-leaved forest

Nitrogen (N) plays a vital role in litter decomposition by interacting with microbial activity in temperate and boreal ecosystems. However, in tropical and subtropical forests, where low soil phosphorus (P) availability is widespread, how P affects litter decomposition remains unclear. In the present study, litter-bag method was used to conduct fine root decomposition by P addition experiment in a subtropical natural evergreen broad-leaved forest. Fine roots were collected from two tree species: Cunninghamia lanceolata and Castanopsis carlesii, and their mixture in equal proportion as well. At the end of 2-year experiment, we found an enhancement of soil total P, available P, pH and microbial biomass carbon content, and increased abundance of all the examined microbial groups and total PLFA biomass after P addition. Annual decomposition constants of fine roots of both tree species and the mixture (0.615–1.049) were all elevated, but to different extents by P addition, with fine roots of lower initial P concentration corresponding to higher increase in decomposition rate. Average acid phosphatase activities during decomposition were significantly decreased, but there was a general increase for β-glucosidase (βG), cellobiohydrolase (CBH), phenol oxidase, peroxidase (PerOx) and N-acetyl glucosaminidase (NAG) activities. The response ratios of enzyme activities, which differed for the three experimental fine roots, corresponded to the individual increase in decomposition rates. Correlation analysis demonstrated that fine root decomposition rates were significantly related to initial P concentration, extractives and acid-insoluble fractions of fine roots, and activities of βG, CBH, NAG and PerOx as well. Taken together, our results demonstrate that fine root decomposition can be constrained by P availability through affecting microbial activity in this subtropical natural forest.

Soil microbial community structure and enzymatic activity along a plant cover gradient in Victoria Land (continental Antarctica)

In continental Antarctica, autotrophs are exclusively represented by cyanobacteria, algae, lichens and mosses. Consequently, Antarctic soil communities are expected to be rather simple and primarily dominated by microorganisms. Recently, a change in abundance of mosses and lichens has been observed in continental Antarctica in response to an increase of the active permafrost layer, but the implication of this change to soil micro-organisms remains little known. Here we aim to clarify to what extent the abundance of mosses and lichens affects soil biogeochemistry in Victoria Land, with a particular focus on soil microbial abundance and associated soil enzymatic activity. To achieve this aim, we assessed the structure of soil microbiome and the activity of hydrolytic C, N, and P enzymes along a gradient in soil physico-chemical conditions and plant cover. Moss cover strongly relates to the amount of soil organic carbon (SOC), soil water and nutrient content. Soils with higher content of organic carbon were characterized by higher microbial biomass and showed a relatively higher abundance of fungi as compared to bacteria. More specifically, PLFAs biomarkers for Actinomycetes and Gram-positive bacteria were mainly associated to soils with lower SOC. In order to sustain a higher microbial biomass, total activity of hydrolytic enzymes increased with increasing SOC content. Eco-enzymatic stoichiometry, based on C to P and C to N ratios, indicates a higher investment in N- and P-hydrolytic enzymes (ratio < 1), particularly at low SOC content. Oppositely, an increase in C-hydrolytic enzyme activity (ratio ≈ 1) was observed with increasing accumulation of organic carbon. Such a result seems to indicate a stronger role of soil pH at low SOC on enzymatic stoichiometry (abiotic control) whereas with increasing accumulation of organic matter the enzymatic stoichiometry is more affected by microbial metabolism (biotic control).

Spatial pattern of enzyme activities depends on root exudate composition

Roots increase microbial activities depending on exudate composition, especially on the ratios of sugars, carboxylic and amino acids, and thus structure enzyme activities in the rhizosphere. We introduce a new approach combining soil zymography and simulated exudates released from Rhizon® samplers to stimulate microbial activities but avoid the direct release of enzymes by living roots. This enabled visualizing, localizing and analyzing the effects of simulated root exudates on activity of five microbial enzymes involved in carbon (C) (β-glucosidase, cellobiohydrolase), nitrogen (N) (leucine aminopeptidase), phosphorus (P) (phosphatase) and sulfur (S) (sulfatase) cycles. We tested the hypotheses that 1) artificial exudates stimulate microorganisms for enzyme production and form spatial gradients around roots, and 2) the extent of microbial enzyme activities in the rhizosphere is component-specific. In line with these hypotheses, the activities of P-, N- and S-related enzymes were higher near the artificial root and gradually decreased as a function of distance from the root. The pattern for C-cycle enzymes was uniform and independent of the exudate composition. Among all components, alanine increased the rhizosphere extent much stronger than other substances, while methionine had no effect on the spatial distribution of enzyme activities. Vmax of all enzymes increased with alanine addition, but decreased after adding citrate. The ratios of enzyme activities demonstrated that rhizosphere microorganisms release more leucine aminopeptidase than other enzymes to meet their N demand. Glucose increased the Km of cellobiohydrolase and β-glucosidase, while alanine had the greatest effect on the Km of leucine aminopeptidase and sulfatase. Phosphatase is the enzyme most sensitive to the composition of root exudates; consequently, any factor influencing root exudate composition can strongly affect the P cycle. We conclude that the rhizosphere extent of microbial-derived enzyme activities is component- and enzyme-specific and that this extent depends on the substrate stoichiometry and microbial nutrient demand.

Temperate grassland shifted from nitrogen to phosphorus limitation induced by degradation and nitrogen deposition: Evidence from soil extracellular enzyme stoichiometry

Soil extracellular enzyme activity (EEA) and enzymatic stoichiometry (ES) can provide a good indication of changes in soil nutrient availability and microbial demands for energy and nutrients. However, across different levels of degraded grasslands, whether the responses of soil EEAs to increasing nitrogen (N) deposition are consistent remains unclear. To explore the effects of N addition on soil EEAs in natural and degraded grasslands, we conducted a 3-year N addition experiment (0, 10, 20, 30, 40 and 50 g N m−2 yr−1) at four sites in temperate grassland of northern China with different levels of degradation. We measured seven soil EEAs, including carbon (C)-acquiring enzyme (β-1,4-glucosidase (BG)), N-acquiring enzymes (β-N-acetyl-glucosaminidase (NAG) and leucine aminopeptidase (LAP)) and phosphorus (P)-acquiring enzyme (acid phosphatase (AP)) activities. The length and angle of vectors defined by ratios of enzyme activities (BG/(NAG + LAP) vs. BG/AP) were used to indicate relative microbial investments in C- (length), and N- and P- (angle) acquiring enzymes. Our results showed that N addition significantly increased AP activity by 176%, 116%, 220% and 268%, decreased LAP activity by 39%, 70%, 67% and 99%, but increased NAG activity by 185%, 137%, 164% and 118% in non-degraded, moderately degraded, severely degraded and extremely degraded grassland, respectively. Decreasing enzyme N:P and C:P ratios and increasing vector angle with the increasing levels of degradation suggested that temperate grassland could transform into P-limitation (angles >45°) from N-limitation (angles <45°). N addition significantly decreased the enzyme N:P ratio and increased the vector angle (angles >45°), indicating that N addition further aggravated P limitation. Our findings highlight that grassland degradation would affect the responses of soil EEAs to N addition and cause nutrient limitation transformation. Soil ES might be a sensitive indicator mediated by soil microorganisms to the relative resource limitation in temperate grasslands.

The engineering and immobilization of penicillin G acylase onto thermo‐sensitive tri‐block copolymer system

In this work, the relationships between catalytic performances of penicillin G acylase (PGA) and the molar ratio of carrier, thermo‐sensitive tri‐block polymer, poly (N,N‐diethylacrylamide‐b‐ β‐hydroxyethyl methacrylate‐b‐glycidyl methacrylate) (PDEA‐b‐PHEMA‐b‐PGMA) were studied firstly, and result documented the optimal molar ratio was nDEA:nHEMA:nGMA = 100:47:24, which presented a suitable lower critical solution temperature (LCST) of 35°C and the activity retention ratio of 80.62% (±0.50%). Based on the suitable carrier, immobilization conditions were investigated and optimized. When pH of solution, concentration of PGA, immobilized time, and immobilization temperature were 8.0, 1/10 (m/v), 16 hours, and 36°C, respectively, enzyme loading capacity (L), enzyme activity (Ea), and activity retention ratio (Ar) of PGA arrived at the highest value of 21 223 U, 16 199 U/g, and 93.50% (±0.50%), respectively. Besides, the response rate (Rr) of immobilized PGA was the same as free PGA, the reusable stability (Rs) was 77.00% (±1.00%) after using for 11 times, which indicated that the carrier has better compatibility with L, Ar, Rs, and Rr.

Litter chemistry influences earthworm effects on soil carbon loss and microbial carbon acquisition

Earthworms could affect soil C and N cycling process to balance their energy and nutrients requirements, and they could also regulate soil microbial community structure and microbial acquisition for C and N. However, the connection between faunal and microbial stoichiometry in the coupling soil C and N cycling remains poorly understood. In a controlled laboratory experiment, we amended soil with five litters differing in litter chemistry (clover, maize stover, wheat straw, Rumex and bagasse fiber) including a no litter control and treated them without or with earthworms (Metaphire guillelmi). After 90 d incubation, we examined changes in earthworm tissue and microbial stoichiometry and different soil C and N fractions. Earthworm tissue C content was rather stable compared with the fluctuation in tissue N, implying that C is under stronger control and associated with higher demand than N. The presence of earthworm significantly enhanced CO2 emissions and decreased particulate organic carbon (POC) and soil organic carbon (SOC) contents in the low lignin litter species clover, maize stover and wheat straw. Meanwhile, earthworm presence increased N2O cumulative emissions but exerted negligible effects on particulate organic nitrogen (PON) and soil total nitrogen (TN) contents irrespective of litter species. Correspondingly, earthworm regulated microbial C and N acquisition as C to N-degrading enzyme activity ratio were nearly doubled in the low lignin litter species clover, maize stover and wheat straw, while it was decreased in the high lignin litter species Rumex and bagasse fiber. However, the structural equation modeling indicated C loss induced by earthworms was mainly attributed to their effects on soil fungi and bacteria abundance, while much less related to C-degrading enzyme activities. In conclusion, litter species controlled earthworm effects on soil C and N loss and associated microbial acquisition for C and N, highlighting the pivotal role of resource chemistry in the regulation of soil fauna impact on soil functioning and ecosystem services.

Bioactive Fungal Screening to find Efficient Biomass‐Degrading Enzymes

Traditional fuels such as coal and gasoline are limited and can produce harmful effects to the environment. Bioethanol from biomass (i.e., plant‐based materials) degradation is a sustainable and renewable form of fuel that aims to reduce these effects. The fungal secretome, known as a pool of biomass degrading enzymes, can be utilized to discover industrial enzyme candidates useful for biofuel production. The overall purpose of our research is to pre‐screen a pool of fungi collected from the Great Lakes, select candidates that can break down biomass (i.e., cellulose) efficiently, and characterize the biomass degrading enzymes. In this study, we first selected seven different fungal species, and grew them onto carboxymethyl cellulose (CMC) plates. A specific dye called Congo red was used to monitor the cellulose activity after either a five‐day or twelve‐day growth period. A clear zone present around the central fungal growth indicated the degradation of the cellulose in the culture medium. Fungal activity was measured using a ratio of enzyme activity (outer ring) and the fungal growth (central circle). The enzyme activities of these strains were compared to Aspergillus niger, a known industrial fungal strain. Overall, there were four different species that demonstrated higher cellulase activity in comparison to A.niger in the CMC assay. Additionally, the secretome and cell lysate of A. niger was prepared to determine the source of the biomass‐degrading enzymes through an enzymatic activity assay. Preparation of the secretome and lysate were obtained through ultra filtration and bead beating, respectively. Both methods resulted in acquiring a highly concentrated solution of each protein samples. It was determined that the biomass‐degrading enzymes were secreted from the fungal cells rather than contained within the cell. Future directions include secretome preparation of the four identified fungal strains to verify their biomass hydrolysis capabilities.Support or Funding InformationThis work was supported NSF award 1359457 to Dr. Si Wu.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Juvenile rockfish show resilience to CO2-acidification and hypoxia across multiple biological scales.

California's coastal ecosystems are forecasted to undergo shifting ocean conditions due to climate change, some of which may negatively impact recreational and commercial fish populations. To understand if fish populations have the capacity to respond to multiple stressors, it is critical to examine interactive effects across multiple biological scales, from cellular metabolism to species interactions. This study examined the effects of CO2-acidification and hypoxia on two naturally co-occurring species, juvenile rockfish (genus Sebastes) and a known predator, cabezon (Scorpaenichthys marmoratus). Fishes were exposed to two PCO2 levels at two dissolved oxygen (DO) levels: ~600 (ambient) and ~1600 (high) μatm PCO2 and 8.0 (normoxic) and 4.5 mg l-1 DO (hypoxic) and assessments of cellular metabolism, prey behavior and predation mortality rates were quantified after 1 and 3 weeks. Physiologically, rockfish showed acute alterations in cellular metabolic enzyme activity after 1 week of acclimation to elevated PCO2 and hypoxia that were not evident in cabezon. Alterations in rockfish energy metabolism were driven by increases in anaerobic LDH activity, and adjustments in enzyme activity ratios of cytochrome c oxidase and citrate synthase and LDH:CS. Correlated changes in rockfish behavior were also apparent after 1 week of acclimation to elevated PCO2 and hypoxia. Exploration behavior increased in rockfish exposed to elevated PCO2 and spatial analysis of activity indicated short-term interference with anti-predator responses. Predation rate after 1 week increased with elevated PCO2; however, no mortality was observed under the multiple-stressor treatment suggesting negative effects on cabezon predators. Most noteworthy, metabolic and behavioral changes were moderately compensated after 3 weeks of acclimation, and predation mortality rates also decreased suggesting that these rockfish may be resilient to changes in environmental stressors predicted by climate models. Linking physiological and behavioral responses to multiple stressors is vital to understand impacts on populations and community dynamics.

Relationship between microbial composition and substrate use efficiency in a tropical soil

Soil organic matter (SOM) is crucial to soil health, supporting most of the soil properties that benefit plant growth and ecosystem services including carbon sequestration, nutrient recycling and water infiltration. Recent study is exposing the soil microbial community as not only decomposing SOM, but also providing the primary source of chemicals for its formation. All else equal, SOM formation is theoretically greatest when microbes maximise enzyme efficiency (ratio of enzyme activity to carbon loss from respiration) and biomass efficiency (biomass gain per unit substrate added). Our study examines the relationship between microbial composition and metrics of these two efficiencies. We hypothesised that both will increase with higher ratios of soil fungi to bacteria. We manipulated microbial composition through sustained use of selective microbial inhibitors in microcosms with tropical soil, alongside variation of litter quality and diversity and the presence or absence of a simulated root exudate. Both litter and inhibitor treatments significantly changed soil microbial composition and function, and enzyme efficiency and biomass efficiency were both higher in microbial communities with more fungi, supporting our hypothesis. Structural equation modelling suggested that the observed efficiency changes did indeed occur in part via changes in microbial composition after accounting for direct effects of treatments. Taken together the results provide some support for the hypothesis that soil fungi benefit SOM formation.

Biochemical proxies indicate differences in soil C cycling induced by long-term tillage and residue management in a tropical agroecosystem

A potential benefit of conservation agriculture (CA) is soil organic carbon (SOC) accrual, yet recent studies indicate limited or no impact of CA on total SOC in tropical agroecosystems. We evaluated biochemical indicators of soil C cycling after 9 years (18 seasons) of contrasting tillage with and without maize residue retention in western Kenya. Potential activities of C-cycling enzymes (β-glucosidase, GLU; β-galactosidase, GAL; glucosaminidase, GLM; cellobiohydrolase, CEL), permanganate-oxidizable C (POXC), and soil organic matter (SOM) composition (by infrared spectroscopy) were measured. POXC tended to be greater under reduced tillage and residue retention, but did not significantly differ among treatments (≤ 2% of SOC). Despite no significant differences in SOC concentrations or stocks, activities of all 4 C-cycling enzymes responded strongly to tillage, and to a lesser extent to residue management. Activities of GLU, GAL, and GLM were greatest under the combination of reduced tillage and residue retention relative to other treatments. Reduced tillage produced an enrichment in carboxyl C = O (+6%) and decreased polysaccharide C-O (−3.5%) relative to conventional tillage irrespective of residue management. Though enzyme activities and POXC are typically associated with SOC accrual, changes in soil C cycling at this site have not translated into significant differences in SOC after 9 years. Elevated enzyme activities may have offset potential SOC accumulation under CA. However, the ratio of C-cycling enzyme activities to SOC was higher under reduced tillage and residue retention relative to other treatments, indicating that stoichiometric scaling of SOC and enzyme activities does not explain absence of significant differences in SOC among tillage and residue managements. Potential factors that may explain the low SOC accrual rates in this tropical agroecosystem included the low, albeit realistic, levels of residue retention, nutrient limitations, and high temperatures favoring decomposition.

Mechanism of resistance to pirimicarb in the cowpea aphid Aphis craccivora

A resistant strain of Aphis craccivora Koch was selected with pirimicarb for 12 generations. The selected strain exhibited 47-fold resistance compared to the susceptible laboratory strain. The resistant strain also showed cross-resistance to carbosulfan, malathion, chlorpyrifos methyl and thiamethoxam. Low resistance to fenitrothion and acetamiprid was also observed. Bioassays in combination with biochemical synergist studies revealed that the higher inhibition of carboxylesterase using the synergist tributyl phosphorotrithioate (DEF) was associated with increased toxicity of pirimicarb in the resistant strain. On the other hand, piperonyl butoxide (PBO) and diethyl maleate (DEM) had less inhibitory effect on mixed function oxidases (MFO) and glutathione-S-transferase (GST) in the resistant strain. The activity of carboxylesterase was 29-fold greater in the resistant strain, whereas the activity of MFO and GST in the resistant strain was only 5.5 and 1.7- fold greater, respectively. The activity of acetylcholine esterase (AChE) in the resistant strain was 2 fold higher than in the susceptible strain. The I50 (the concentration of pirimicarb that inhibit 50% of the enzyme activity) ratio of R-strain to S-strain was 12.6. Molecular studies using real-time quantitative PCR showed that the transcription level of Ace2 gene in the resistant strain was 3.4-fold higher than that in the susceptible strain. In conclusion, mechanism of resistance in the pirimicarb resistant strain of A. craccivora may include overexpression of Ace2 gene and higher activity of the detoxification enzymes esterases and partly MFO.

Possible involvement of phosphoenolpyruvate carboxylase and NAD-malic enzyme in response to drought stress. A case study: a succulent nature of the C4-NAD-ME type desert plant, Salsola lanata (Chenopodiaceae).

The co-ordination between the primary carboxylating enzyme phosphoenolpyruvate carboxylase (PEPC) and the further decarboxylating enzymes is crucial to the efficiency of the CO2-concentrating mechanism in C4 plants, and investigations on more types of C4 plants are needed to fully understand their adaptation mechanisms. In this study we investigated the effect of drought on carboxylating enzyme PEPC, and the further decarboxylating NAD-malic enzyme (NAD-ME) of Salsola lanata Pall. (Chenopodiaceae) - an annual succulent C4-NAD-ME subtype desert plant. We investigated enzyme activity at the transcriptional level with real-time quantitative PCR and at the translational level by immunochemical methods, and compared S. lanata with other forms of studied C4 plants under drought stress. Results showed that only severe stress limited PEPC enzyme activity (at pH 8.0) of S. lanata significantly. Considering that PEPC enzyme activity (at pH 8.0) was not significantly affected by phosphorylation, the decrease of PEPC enzyme activity (at pH 8.0) of S. lanata under severe stress may be related with decreased PEPC mRNA. The suggestion of increased phosphorylation of the PEPC enzyme in plants under moderate stress was supported by the ratio of PEPC enzyme activity at pH 7.3/8.0, as PEPC enzyme is inhibited by L-malate and the evidence of the 50% inhibiting concentration of L-malate. NAD-ME activity decreased significantly under moderate and severe stress, and coincided with a change of leaf water content rather than the amount of α-NAD-ME mRNA and protein. Leaf dehydration may cause the decrease of NAD-ME activity under water stress. Compared with other C4 plants, the activities of PEPC and NAD-ME of S. lanata under drought stress showed distinct features.

Detoxifying enzyme studies on cotton leafhopper, Amrasca biguttula biguttula (Ishida), resistance to neonicotinoid insecticides in field populations in Karnataka, India

AbstractThe cotton leafhopper (Amrasca biguttula biguttula Ishida) is considered to be an alarming insect pest causing both quantitative and qualitative loss in cotton. In situ bioassay studies were done and the role of detoxifying enzymes in conferring resistance to neonicotinoid groups of insecticides in low (MUD), medium (DVG), high (HVR) and very high (GLB) pesticide usage areas of Karnataka were determined. Bioassay studies showed that imidacloprid, thiamethoxam, acetamiprid, thiacloprid and clothianidin registered varying levels of resistance for all the locations studied. The resistance ratio was high in imidacloprid (3.35, 8.57, 9.15 and 12.27 fold respectively) and the lowest in dinoferuran (1.86, 5.13, 6.71 and 9.88 fold respectively). Furthermore, the enzyme activity ratio (glutathione-S-transferase) was relatively greater, and corresponded to the higher LC50values of neonicotinoids for very high, high, medium and low pesticide usage areas. Our study suggested that the higher activity of the detoxifying enzyme in the resistance population of cotton leafhopper apparently has a significant role in endowing resistance to neonicotinoid groups of insecticides. However, this study recommends using neonicotinoids in cotton growing areas with caution.

English

The increase in oxidative enzyme activities is related to diminished mite infestation. Some biological aspects of Raoiella indica Hirst reared on the coconut cultivars (‘Jamaican Tall’ (JT), ‘Malayan Yellow Dwarf’ (MYD), Niu Leka (NL) and a hybrid JT x MYD) were studied under laboratory conditions. Aditionally, changes in oxidative enzyme activities (peroxidase and polyphenol oxidase) as response to R. indica feeding were studied in the cultivars where red palm mites showed highest and lowest biological parameters values. Longer time spans and lower oviposition rates observed on the JT suggest this cultivar to be more resistant to R. indica feeding. Cultivar JT showed the highest value in PPO/POX ratio, being about twice the value shown by MYD in the infested plants. The observed enzyme activity ratios in both genotypes showed a slight increase 24 h after mite infestation, suggesting these enzymes could be related to plant resistance to R. indica. However, this relationship is still unclear. The biological parameters of R. indica together with higher enzyme activity, particularly on JT suggest this cultivar could be considered as a more resistant cultivar as compared to MYD. More detailed studies are required to determine the effect of these enzymes on coconut resistance to red palm mites. Key words: Coconut, peroxidase, polyphenol oxidase, red palm mite.

The altitudinal effects on photosynthesis of Rosa platyacantha from the Tianshan Mountains in Northwestern China

The altitudinal effects on photosynthesis were measured on progenies of three populations of Rosa platyacantha Schrenk from altitudes of 1,170 (L); 1,580 (M); and 1,920 (H) m a. s. l. During the day, net photosynthetic rate (P N) decreased in all populations due to the high air temperature in the summer. The H population showed a significantly lower P N at noon compared to other populations. The midday depression of P N occurred in L and M populations due to stomatal limitations, while P N inhibition was associated with PSII activity decline in the H population. In order to avoid photodamage, the plants of H population triggered active antioxidant defenses with a higher enzyme activity and redox ratio of ascorbate at midday compared to the L and M populations. However, more oxidative injury still occurred in the H plants at noon due to higher lipid peroxidation. Our results indicated that the provenance significantly affected photosynthesis in R. platyacantha from northwestern China.

Optimisation of the synthesis of high galacto-oligosaccharides (GOS) from lactose with β-galactosidase from Kluyveromyces lactis

A rational optimisation for the synthesis of galacto-oligosaccharides (GOS) catalysed by a commercial β-galactosidase from Kluyveromyces lactis, Lactozym Pure 6500 L, is shown. The study of the main reaction parameters – temperature, enzyme concentration, pH, initial lactose concentration and reaction time – using surface response methodology, was demonstrated to be an accurate tool to optimise empirical production of the most desired galacto-oligosaccharides (tri-GOS and tetra-GOS) with a higher presumed prebiotic effect. The optimal value for the yield towards these high-GOS predicted by the model was 12.18% at 40 °C, 5 U mL−1 enzyme concentration, pH 7.0, 250 g L−1 initial lactose concentration and 3 h of reaction. The pH was found to be a critical parameter affecting the transgalactosylation/hydrolysis enzyme activity ratio, and was used to tune the relative production of tri- or tetra-GOS.

Stoichiometry of soil extracellular enzyme activity along a climatic transect in temperate grasslands of northern China

Ratios of specific carbon (C), nitrogen (N) and phosphorus (P) acquisition activities converged on 1:1:1 at a global scale and in tropical ecosystems. It is less clear if this pattern can be applied in temperate grasslands. The questions of whether the pattern remains stable across different soil depths and what the relative contributions are from the influence of climatic, edaphic abiotic and biotic factors on soil extracellular enzyme activity (EEA) stoichiometry remain uncertain. We measured potential activities of one C-acquiring enzyme (β-1, 4-glucosidase), two N-acquiring enzymes (β-N-acetylglucosaminidase and leucine aminopeptidase) and one organic P-acquiring enzyme (acid phosphatase) and major influential factors along a climatic transect in temperate grasslands of northern China during the growing season of 2013. We found lower enzyme C: N (0.47) and C: P (0.18) ratios and a higher enzyme N: P activity ratio (0.40) in 0–20 cm soil depth in temperate grasslands than in tropical soils (C: N, 1.83; C:P, 0.21; N: P, 0.13). The enzyme C: N and C: P ratios decreased with soil depth except for the enzyme C: N ratio in desert steppes. However, there were no significant differences in enzyme N: P ratio with soil depth. Among all the factors, soil total C, N and P contents accounted for the most variation in soil EEA and ecoenzymatic stoichiometry in 0–10 cm surface soil, which implied that soil EEA stoichiometry was largely controlled by soil nutrient stoichiometry. Moreover, edaphic factors had less influence on soil EEA stoichiometry in subsoil than in surface soil and edaphic abiotic factors had a larger effect on soil EEA stoichiometry than climatic and biotic factors. Our results suggest that soil extracellular enzyme activity ratios were not in homeostasis but resource dependent on soil and microbial biomass stoichiometry.