Conformational change in the activation of lipase: an analysis in terms of low-frequency normal modes.

The interfacial activation of Rhizomucor miehei lipase (RmL) involves the motion of an alpha-helical region (residues 82-96) which acts as a "lid" over the active site of the enzyme, undergoing a displacement from a "closed" to an "open" conformation upon binding of substrate. Normal mode analyses performed in both low and high dielectric media reveal that low-frequency vibrational modes contribute significantly to the conformational transition between the closed and open conformations. In these modes, the lid displacement is coupled to local motions of active site loops as well as global breathing motions. Atomic fluctuations of the first hinge of the lid (residues 83-84) are substantially larger in the low dielectric medium than in the high dielectric medium. Our results also suggest that electrostatic interactions of Arg86 play an important role in terms of both the intrinsic stability of the lid and its displacement, through enhancement of hinge mobility in a high dielectric medium. Additional calculations demonstrate that the observed patterns of atomic fluctuations are an intrinsic feature of the protein structure and not dependent on the nature of specific energy minima.     

Year-round observations of the net ecosystem exchange of carbon dioxide in a native tallgrass prairie

An Ameriflux site was established in mid 1996 to study the exchange of CO₂ in a native tallgrass prairie of north‐central Oklahoma, USA. Approximately the first 20 months of measurements (using eddy covariance) are described here. This prairie, dominated by warm season C4 grasses, is typical of the central Kansas/northern Oklahoma region. During the first three weeks of the measurement period (mid‐July–early August 1996), moisture‐stress conditions prevailed. For the remainder of the period (until March 1998), however, soil moisture was nonlimiting. Mid‐day net ecosystem CO₂ exchange (NEE), under well‐watered conditions, reached a maximum magnitude of 1.4 mg CO₂ m^?2 s^?1 (flux toward the surface is positive) during peak growth (mid‐July 1997), with green leaf area index of 2.8. In contrast, under moisture‐stress conditions in the same growth stage in 1996, mid‐day NEE was reduced to near‐zero. Average night NEE ranged from near‐zero, during winter dormancy, to ? 0.50 mg CO₂ m^?2 s^?1, during peak growth. Most of the variance in average night NEE was explained by changes in soil temperature (0.1 m depth) and green leaf area. The daytime NEE measurements were examined in terms of a rectangular hyperbolic relationship with incident photosynthetically active radiation. The analysis showed that the quantum yield during peak growth was similar to those measured in other prairies and the y‐intercept, so obtained, can be potentially used as an estimate of night‐time CO₂ emissions when eddy covariance data are unavailable. Daily integrated NEE reached its peak magnitude of 30.8 g CO₂ m^?2 d^?1 (8.4 g C m^?2 d^?1) in mid‐July when the green LAI was the largest (about 2.8). In general, the seasonal trend of daily NEE (on relatively clear days) followed that of green LAI. Annually integrated carbon exchange, between prescribed burns in 1997 and 1998, was 268 g C m^?2 y^?1. After incorporating carbon loss during the prescribed burn , the net annual carbon exchange in this prairie was near‐zero in 1998.     

Tumor cell imaging using the intrinsic emission from PAMAM dendrimer: a case study with HeLa cells

HeLa 229 cells were treated with methotrexate (MTX) and doxorubicin (DOX), utilizing fourth generation (G4), amine terminated poly(amidoamine) {PAMAM} dendrimer as the drug carrier. In vitro kinetic studies of the release of both MTX and DOX in presence and absence of G4, amine terminated PAMAM dendrimers suggest that controlled drug release can be achieved in presence of the dendrimers. The cytotoxicity studies indicated improved cell death by dendrimer-drug combination, compared to the control experiments with dendrimer or drug alone at identical experimental conditions. Furthermore, HeLa 229 cells were imaged for the first time utilizing the intrinsic emission from the PAMAM dendrimers and drugs, without incorporating any conventional fluorophores. Experimental results collectively suggest that the decreased rate of drug efflux in presence of relatively large sized PAMAM dendrimers generates high local concentration of the dendrimer-drug combination inside the cell, which renders an easy way to image cell lines utilizing the intrinsic emission properties of PAMAM dendrimer and encapsulated drug molecule.     

Involvement of Lipid-linked Oligosaccharides in Synthesis of Storage Glycoproteins in Soybean Seeds

Membrane preparations from developing soybean (var. Prize) cotyledon tissue, at the time of synthesis of storage glycoproteins, catalyze the sequential assembly of lipid-linked oligosaccharides from uridine-5'-diphospho-N-acetyl-d-[6-(3)H] glucosamine and guanosine-5'diphospho-d-[U-(14)C]mannose. The maximum size of lipid-linked oligosaccharide that accumulates contains the equivalent of 10 saccharide units on the basis of Bio-Gel P-2 gel filtration studies. These lipid-linked oligosaccharides show similar characteristics to polyisoprenyl diphosphate derivatives on diethylaminoethyl-cellulose chromatography and are potential intermediates in glycoprotein biosynthesis in this tissue. These glycolipids do not appear to turn over in pulse-chase experiments and no completed storage glycoproteins were detected among the products of these incubations.Tissue slices from cotyledons at the same stage of development synthesize lipid-linked oligosaccharides from [(3)H]mannose and [(3)H]glucosamine with sizes equivalent to 1, 7, 10, and approximately 15 saccharide units. In pulse-chase experiments, the lipid-linked saccharides with the equivalent of 1 and 10 units rapidly turnover, whereas those with 7 and 15 units do not. Examination of the higher oligosaccharide peaks (10 and 15) by Bio-Gel P-4 gel filtration shows them to comprise 2 distinct subsets of oligosaccharides containing different proportions of glucosamine and mannose units. Tissue slices synthesize products which resemble the completed 7S storage glycoproteins as judged by similarity of molecular weight and precipitation with specific antisera. Analysis of the oligosaccharides obtained by hydrazinolysis of glycoproteins shows the presence of a similar size "high-mannose" type N-linked oligosaccharides as in other glycoproteins from animal and plant cells.     

DNA biosensor for detection of Neisseria gonorrhoeae causing sexually transmitted disease

A sequence-specific electrochemical sexually transmitted disease (STD) sensor based on self-assembled monolayer of thiolated DNA probe specific to target opa gene for detection of Gonorrhoea, a sexually transmitted disease has been fabricated. 6-Mercapto-1-hexanol (MCH) has been used as a blocking agent to facilitate oligos "stand" up at the surface, a configuration favoring subsequent DNA hybridization and to repel non-specific adsorption of undesired DNA. The results of differential pulse voltammetric studies of this STD sensor reveal low detection limit (1.0 × 10(-18)M) and a wide dynamic range (from 1.0 × 10(-6)M to 0.5 × 10(-18)M) arising due to the stable hybridization using methylene blue as an electro-active DNA hybridization indicator. The experimental results with genomic DNA, clinical patient sample of Neisseria gonorrhoeae, culture of non-N. gonorrhoeae Neisseria species (NgNS) and gram negative bacteria indicate that the fabricated sensor is specific to this STD.     

Surface trapping and AFM detection of DNA topological intermediates generated from an oxidative chemical nuclease

Direct observation of DNA topological intermediates generated from a 'chemical nuclease' treatment has been made by atomic force microscopy (AFM). The intermediates were trapped at the mica-water interface and imaging was carried out in the dynamic force mode. Complete conversion from supercoiled circular state to relaxed circular/linear state has been observed over a time scale of 8 min. Implication of such studies in complementing gel electrophoresis data has been predicted.     

Pesticide relevance and their microbial degradation: a-state-of-art

The extensive use of pesticide causes imbalance in properties of soil, water and air environments due to having problem of natural degradation. Such chemicals create diverse environmental problem via biomagnifications. Currently, microbial degradation is one of the important techniques for amputation and degradation of pesticide from agricultural soils. Some studies have reported that the genetically modified microorganism has ability to degrade specific pesticide but problem is that they cannot introduce in the field because they cause some other environmental problems. Only combined microbial consortia of indigenous and naturally occurring microbes isolated from particular contaminated environment have ability to degrade pesticides at faster rate. The bioaugumentation processes like addition of necessary nutrients or organic matter are required to speed up the rate of degradation of a contaminant by the indigenous microbes. The use of indigenous microbial strains having plant growth activities is ecologically superior over the chemical methods. In this review, we have attempted to discuss the recent challenge of pesticide problem in soil environment and their biodegradation with the help of effective indigenous pesticides degrading microorganisms. Further, we highlighted and explored the molecular mechanism for the pesticide degradation in soil with effective indigenous microbial consortium. This review suggests that the use of pesticide degrading microbial consortia which is an eco-friendly technology may be suitable for the sustainable agriculture production.