Role of Arg-401 of cytosolic serine hydroxymethyltransferase in subunit assembly and interaction with the substrate carboxy group.

In an attempt to identify the arginine residue involved in binding of the carboxylate group of serine to mammalian serine hydroxymethyltransferase, a highly conserved Arg-401 was mutated to Ala by site-directed mutagenesis. The mutant enzyme had a characteristic visible absorbance at 425 nm indicative of the presence of bound pyridoxal 5'-phosphate as an internal aldimine with a lysine residue. However, it had only 0.003% of the catalytic activity of the wild-type enzyme. It was also unable to perform reactions with glycine, beta-phenylserine or d-alanine, suggesting that the binding of these substrates to the mutant enzyme was affected. This was also evident from the interaction of amino-oxyacetic acid, which was very slow (8.4x10(-4) s-1 at 50 microM) for the R401A mutant enzyme compared with the wild-type enzyme (44.6 s-1 at 50 microM). In contrast, methoxyamine (which lacks the carboxy group) reacted with the mutant enzyme (1.72 s-1 at 250 microM) more rapidly than the wild-type enzyme (0.2 s-1 at 250 microM). Further, both wild-type and the mutant enzymes were capable of forming unique quinonoid intermediates absorbing at 440 and 464 nm on interaction with thiosemicarbazide, which also does not have a carboxy group. These results implicate Arg-401 in the binding of the substrate carboxy group. In addition, gel-filtration profiles of the apoenzyme and the reconstituted holoenzyme of R401A and the wild-type enzyme showed that the mutant enzyme remained in a tetrameric form even when the cofactor had been removed. However, the wild-type enzyme underwent partial dissociation to a dimer, suggesting that the oligomeric structure was rendered more stable by the mutation of Arg-401. The increased stability of the mutant enzyme was also reflected in the higher apparent melting temperature (Tm) (61 degrees C) than that of the wild-type enzyme (56 degrees C). The addition of serine or serinamide did not change the apparent Tm of R401A mutant enzyme. These results suggest that the mutant enzyme might be in a permanently 'open' form and the increased apparent Tm could be due to enhanced subunit interactions.

Role of cell attachment in leaching of chalcopyrite mineral by Thiobacillus ferrooxidans

Experiments on the leaching of copper from chalcopyrite mineral by the bacterium Thiobacillus ferrooxidans show that, in the presence of adequate amounts of sulphide, iron-grown bacteria preferentially oxidise sulphur in the ore (through direct attachment) rather than ferrous sulphate in solution. At 20% pulp density, the leaching initially takes place by a predominantly direct mechanism. The cell density in the liquid phase increases, but the Fe2+ is not oxidised. However, in the later stages when less solid substrate is available and the cell density becomes very high, the bacteria start oxidising Fe2+ in the liquid phase, thus contributing to the indirect mechanism of leaching. Contrary to expectations, the rate of leaching increased with increasing particle size in spite of the decreasing specific surface area. This has been found to be due to increasing attachment efficiency with increase in particle size.

A Demonstration of the Role ofhetGenes in Heterokaryon Formation inNeurosporaunder Simulated Field Conditions

An experimental system was developed for assessing the role ofhetgenes in heterokaryon formation inNeurosporain nature. Burned sugar cane segments planted in soil were infected using a mixture of mutant ascospores of two genotypes.Neurosporaramified in the cane and erupted as distinct pustules of conidia. When ascospores carried identicalhetalleles, the (macro) conidial pustules which formed were heterokaryotic. On the other hand, when ascospores carried dissimilarhetalleles, the pustules were homokaryotic. These results showed that stable heterokaryons between compatible strains can form in nature. When two strains are growing together on a natural substrate, heterozygosity athetloci serves to maintain their individuality.

Role of Thiobacillus ferrooxidans and sulphur (sulphide)-dependent ferric-ion-reducing activity in the oxidation of sulphide minerals

Thiobacillus ferrooxidans oxidized the sulphide minerals e.g., pyrite, pyrrhotite and copper concentrate under anaerobic conditions in the presence of ferric ion as sole electron acceptor. Copper and iron were solubilized from sulphide ores by the sulphur (sulphide)-dependent ferric-ion oxidoreductase activity. Treatment of resting cells of T. ferrooxidans with 0.5% phenol for 30 min completely destroyed the iron- and copper-solubilizing activity. The above treatment destroyed the sulphur(sulphide)-dependent ferric-ion-reducing activity completely but did not affect the iron-oxidizing activity. The results suggest that sulphur(sulphide)-dependent ferric-ion-reducing activity actively participates in the oxidation of sulphide minerals under anaerobic conditions. The activity of sulphur(sulphide)-dependent ferric ion reduction in the solubilization of iron and copper from the sulphide ores were also observed under aerobic conditions in presence of sodium azide (0.1 μmol), which completely inhibits the iron-oxidizing activity.

Why and when do learning goal orientation and attitude decrease with aging? The role of perceived remaining time and work centrality

We conducted two studies to improve our understanding of why and when older workers are focused on learning. Based on socioemotional selectivity theory, which proposes that goal focus changes with age and the perception of time, we hypothesized and found that older workers perceive their remaining time at work as more limited than younger workers which, in turn, is associated with lower learning goal orientation and a less positive attitude toward learning and development. Furthermore, we hypothesized and found that high work centrality buffers the negative association between age and perceived remaining time, and thus the indirect negative effects of age on learning goal orientation and attitude toward learning and development (through perceived remaining time). These findings suggest that scholars and practitioners should take workers’ perceived remaining time and work centrality into account when examining or stimulating learning activities among aging workers.

The role of lakes in carbon cycling in boreal catchments

Lakes are an important component of ecosystem carbon cycle through both organic carbon sequestration and carbon dioxide and methane emissions, although they cover only a small fraction of the Earth's surface area. Lake sediments are considered to be one of rather perma-nent sinks of carbon in boreal regions and furthermore, freshwater ecosystems process large amounts of carbon originating from terrestrial sources. These carbon fluxes are highly uncer-tain especially in the changing climate. -- The present study provides a large-scale view on carbon sources and fluxes in boreal lakes situated in different landscapes. We present carbon concentrations in water, pools in lake se-diments, and carbon gas (CO2 and CH4) fluxes from lakes. The study is based on spatially extensive and randomly selected Nordic Lake Survey (NLS) database with 874 lakes. The large database allows the identification of the various factors (lake size, climate, and catchment land use) determining lake water carbon concentrations, pools and gas fluxes in different types of lakes along a latitudinal gradient from 60oN to 69oN. Lakes in different landscapes vary in their carbon quantity and quality. Carbon (C) content (total organic and inorganic carbon) in lakes is highest in agriculture and peatland dominated areas. In peatland rich areas organic carbon dominated in lakes but in agricultural areas both organic and inorganic C concentrations were high. Total inorganic carbon in the lake water was strongly dependent on the bedrock and soil quality in the catchment, especially in areas where human influence in the catchment is low. In inhabited areas both agriculture and habitation in the catchment increase lake TIC concentrations, since in the disturbed soils both weathering and leaching are presumably more efficient than in pristine areas. TOC concentrations in lakes were related to either catchment sources, mainly peatlands, or to retention in the upper watercourses. Retention as a regulator of the TOC concentrations dominated in southern Finland, whereas the peatland sources were important in northern Finland. The homogeneous land use in the north and the restricted catchment sources of TOC contribute to the close relationship between peatlands and the TOC concentrations in the northern lakes. In southern Finland the more favorable climate for degradation and the multiple sources of TOC in the mixed land use highlight the importance of retention. Carbon processing was intensive in the small lakes. Both CO2 emission and the Holocene C pool in sediments per square meter of the lake area were highest in the smallest lakes. How-ever, because the total area of the small lakes on the areal level is limited, the large lakes are important units in C processing in the landscape. Both CO2 and CH4 concentrations and emissions were high in eutrophic lakes. High availability of nutrients and the fresh organic matter enhance degradation in these lakes. Eutrophic lakes are often small and shallow, enabling high contact between the water column and the sediment. At the landscape level, the lakes in agricultural areas are often eutrophic due to fertile soils and fertilization of the catchments, and therefore they also showed the highest CO2 and CH4 concentrations. Export from the catchments and in-lake degradation were suggested to be equally important sources of CO2 and CH4 in fall when the lake water column was intensively mixed and the transport of sub-stances from the catchment was high due to the rainy season. In the stagnant periods, especially in the winter, in-lake degradation as a gas source was highlighted due to minimal mixing and limited transport of C from the catchment. The strong relationship between the annual CO2 level of lakes and the annual precipitation suggests that climate change can have a major impact on C cycling in the catchments. Increase in precipitation enhances DOC export from the catchments and leads to increasing greenhouse gas emissions from lakes. The total annual CO2 emission from Finnish lakes was estimated to be 1400 Gg C a-1. The total lake sediment C pool in Finland was estimated to be 0.62 Pg, giving an annual sink in Finnish lakes of 65 Gg C a-1.