TheTy1-copia group retrotransposons inVicia species: copy number,sequence heterogeneity and chromosomal localisation

We present an in-depth study of theTy1-copia group of retrotransposons within the plant genusVicia, which contains species with widely differing genome sizes. We have compared the numbers and sequence heterogeneities of these genetic elements in three diploidVicia species chosen to represent large (V. faba, 1C=13.3 pg), medium (V. melanops, 1C=11.5 pg) and small (V. sativa, 1C=2.3 pg) genomes within the genus. The copy numbers of the retrotransposons are all high but vary greatly, withV. faba containing approximately 10⁶ copies,V. melanops about 1000 copies andV. sativa 5000 copies. The degree of sequence heterogeneity ofTy1-copia group elements correlates with their copy number within each genome, but neither heterogeneity nor copy number are related to the genome size of the host. In situ hybridization to metaphase chromosomes shows that the retrotransposons inV. faba are distributed throughout all chromosomes but are much less abundant in certain heterochromatic regions. These results are discussed in the context of plant retrotransposon evolution.     

Melatonin Does Not Modulate Testicular Responsiveness to Altered Photoperiods. A Study During Different Phases of the Annual Gonadal Cycle in Roseringed Parakeets (Psittacula krameri)

The roseringed parakeet has been shown to exhibit a variable testicular responsiveness to both altered photoperiodic regimens and to treatment with melatonin during different phases of the annual gonadal cycle. Adult male roseringed parakeets were held under either natural photoperiods (NP), or long photoperiods (LP; 16L 8D), or short photoperiods (SP; 8L 16D) for a total period of 90 days. From day 46 onward, half of the total birds in each group were administered with the vehicle of melatonin, and the other birds were injected daily in the afternoon with melatonin (25 µg/ 100 g body wt.) till the end of the experiment. An identical experimental schedule was followed during the four different (preparatory, progressive, pre-breeding, and breeding) phases of the annual testicular cycle. The testicular activities in various bird groups were evaluated by volumetric, gravimetric, histometric and karyometric measurements, and by quantitative histological studies. The findings revealed that exogenous melatonin may exert either a suppressive influence or none at all on the testicular functions in relation to the photoperiodic schedule as well as to the reproductive phase of the concerned bird, but in no case modulates gonadal responsiveness to artificially altered photoperiods.     

Role of some riparian wetland plants in reducing erosion of organic carbon and selected cations

Five riparian herbaceous plants, Leonotis nepetaefolia, Cassia tora, Ageratum conyzoides, Parthenium hysterophorus and Sida acuta, dominant on the banks of the Rihand river at Renukoot (India), were selected to assess experimentally their quantitative role in conserving organic-C, Na, K and Ca. Young seedlings from the river bank were planted on sloping experimental plots made of alluvial soil. Simulated rainfall totalling 42.5 mm was applied at 300 mm h⁻¹ on five vegetated and one bare plots. Runoff water and eroded soil were collected from each experimental plot in artificial reservoirs and their quantities were measured. The soil conservation value of the five selected species ranged between 33 and 84% while the water conservation value varied between 19 and 50%. The overall nutrient conservation value, based on the losses in runoff water and eroded soil taken together, varied from 30 to 83% for organic-C, 19 to 78% for Na, 13 to 72% for K and 29 to 52% for Ca under different species. Loss of these four nutrients in response to 42.5 mm simulated rainfall was much higher than their input through rainfall. Loss value for the nutrients were in following order: organic-C > Ca > K > Na. The fraction of organic-C transported down the slope was higher in eroded soil (averaging 73%) and of exchangeable bases in runoff water (averaging 86% for Na, 82% for K and 90% for Ca). Flow-weighted concentrations of all the studied nutrients were consistently greater from bare stands. Number of fine roots was found to play greater role in the case of organic-C (92%; p < 0.01) and Na (70%; p < 0.05) runoff and their conservation by different plant species but canopy cover played greater role for K (58%; p < 0.08) and Ca (90%; p < 0.01).     

Chimeric hemoglobins--hybrids of human and swine hemoglobin: assembly and stability of interspecies hybrids.

Transgenic swine expressing human HbA contained only one of two types of the anticipated interspecies hybrids, namely H alpha 2 P beta 2 (H = human, P = swine). In an attempt to establish whether the absence of the swine alpha and human beta (P alpha 2 H beta 2) hybrid in vivo is a reflection of the lack of complementarity between the interspecies chains to generate appropriate interfaces, we have undertaken the in vitro assembly of swine alpha and human beta chimeric tetramer. In contrast to the in vivo transgenic swine system, in vitro the hybrid of swine alpha human beta chain is assembled readily and the hybrid exhibits normal cooperative oxygen binding. Both the swine alpha human beta and the human alpha swine beta interspecies hybrids are stable around neutral pH and do not segregate into parent tetramers even when mixed together. On the other hand, nearly complete exchange of P alpha chain of P alpha 2 H beta 2 hybrid occurs in the presence of H alpha chain at pH 6.0 and room temperature, resulting in the formation of HbA. However, very little of such an exchange reaction takes place at pH 7.0. These results suggest that the thermodynamic stability of P alpha 2 H beta 2 hybrid is lower compared to that of HbA. In contrast, P beta chain of H alpha 2 P beta 2 hybrid is refractory to exchange with H beta chain at pH 7.0 as well as at pH 6.0, suggesting that the stability of H alpha 2 P beta 2 is higher compared to that of HbA (H alpha 2 H beta 2). The swine alpha human beta chimeric Hb undergoes subunit exchange reaction with human alpha-chain in the presence of 0.9 M MgCl2, at pH 7.0. This demonstrates the lower thermodynamic stability of the intradimeric interactions of the heterodimer even at neutral pH. A synergistic coupling of the intra- and interdimeric interactions of the swine alpha and human beta chain heterodimer is essential for the thermodynamic stability of the chimeric Hb under the physiological conditions. Accordingly, we speculate that the lower thermodynamic stability of P alpha H beta heterodimer (compared to the homodimers H alpha H beta and P alpha P beta) facilitates its segregation into the homodimers by subunit exchange reaction involving either H alpha or P beta. This molecular aspect by itself or possibly along with other cellular aspects of the swine system results in the absence of P alpha 2 H beta 2 hybrid in transgenic swine expressing HbA.     

Production, Purification, and Characterization of Recombinant Human Hemoglobin Rainier Expressed inSaccharomyces cerevisiae

Hemoglobin Rainier is a naturally occurring hemoglobin variant in which the β145 tyrosine is substituted with cysteine. The α and β^Rainierglobin cDNAs were cloned in a high copy number vector and expressed inSaccharomyces cerevisiaeunder the control of galactose-regulated hybrid promoters. Using this system, we have expressed individual α and β^Rainierglobin chains. Coexpression of both α and β^RainiercDNAs resulted in the production of a functional hemoglobin molecule. Purification of the recombinant protein was accomplished by ion exchange chromatography. The N-termini of the α and β chains were correctly processed, and the molecular mass, as determined by mass spectrometry, indicated amino acid composition identical to that of natural hemoglobin Rainier. The chromatographic properties of the recombinant hemoglobin Rainier were similar to human-derived hemoglobin A₀. The purified recombinant hemoglobin molecule was shown to have an elevated oxygen affinity and a reduced cooperativity as previously reported for natural hemoglobin Rainier. Production of recombinant hemoglobin and especially hemoglobin variants like hemoglobin Rainier has the potential to facilitate use of hemoglobin as a blood substitute as well as in specific applications, such as for use as a therapeutic agent in the treatment of hypotension associated with septic shock.     

Cell-Free Synthesis of Pectin (Identification and Partial Characterization of Polygalacturonate 4-[alpha]-Galacturonosyltransferase and Its Products from Membrane Preparations of Tobacco Cell-Suspension Cultures)

Polygalacturonate 4-[alpha]-galacturonosyltransferase (EC 2.4.1.43) activity has been identified in microsomal membranes isolated from tobacco (Nicotiana tabacum L. cv Samsun) cell-suspension cultures. Incubation of UDP-[14C]galacturonic acid with tobacco membranes results in a time-dependent incorporation of [14C]galacturonic acid into a chloroform-methanol-precipitable and 65% ethanol-insoluble product. The optimal synthesis of product occurs at a pH of 7.8, 25 to 30[deg]C, an apparent Km for UDP-D-galacturonic acid of approximately 8.9 [mu]M, and a Vmax of approximately 150 pmol min-1 mg-1 protein. The product was characterized by scintillation counting, thin-layer chromatography, high-performance anion-exchange chromatography, and gel-filtration chromatography in combination with enzymatic and chemical treatments. The intact product has a molecular mass of approximately 105,000 D based on dextran molecular standards. The product was treated with base to hydrolyze ester linkages (e.g. methyl esters), digested with a homogeneous endopolygalacturonase (EPGase), or base and EPGase treated. Base and EPGase treatment results in cleavage of 34 to 89% of 14C-labeled product into components that co-chromatograph with mono-, di-, and trigalacturonic acid, indicating that a large portion of product contains contiguous 1,4-linked [alpha]-D-galactosyluronic acid residues. Optimal EPGase fragmentation of the product requires base treatment prior to enzymatic digestion, suggesting that 45 to 67% of the galacturonic acid residues in the synthesized homogalacturonan are esterified. At least 40% of the base-sensitive linkages were shown to be methyl esters by comparing the sensitivity of base-treated and pectin methylesterase-treated products to fragmentation by EPGase.     

Ethanol-mediated gene-amplification in Escherichia coli enhances derepressed synthesis of β-galactosidase

Summary The presence of ethanol (5 % v/v), in nutrient medium, ehanced DNA synthesis per E. coli cell nearly 2.8-fold compared to that in control cells. At this concentration, the derepressed synthesis of -galactosidase per bacterium also increased about 3-fold. We, therefore, propose that the ethanol-mediated gene-amplification proportionately elevated the induced synthesis of -galactosidase.     

A novel cyclomaltodextrin glucanotransferase from Bacillus firmus that degrades raw starch

Summary An alkalophilic Bacillus firmus secreting the enzyme cyclomaltodextrin glucanotransferase was isolated from soil. The enzyme attacked raw starch to produce cyclodextrins. Maximum cyclodextrins were produced from tapioca starch followed by potato and corn starch. About 49 % of tapioca starch (at 10 and 50 g/l) was converted to cyclodextrins. The main reaction products were and -cyclodextrins with 40 % and 8 % yield respectively. On prolonged incubation small amount of -cyclodextrin was also produced. The ratio of cyclodextrins was dependent on the initial substrate concentration as well as reaction time.NCL communication number 6203     

Citric acid fermentation by the mutant strain of theAspergillus niger resistant to manganese ions inhibition

Summary A new mutant strain,Aspergillus niger GS-III, showing resistance to manganese ions inhibition of citric acid fermentation on a sugarcane molasses containing medium was induced fromAspergillus niger KCU 520, a high citric acid-yielding strain. In submerged, surface or continuous cultures in the presence of manganese ions concentration upto 1.5 ppm the mutant strain yielded citric acid about 90 KgM^–3 . The citric acid yield was comparable to that obtained with the parental strain KCU 520 in the absence of manganese ions, but it was atleast 3-fold higher than that obtained by the latter in the presence of manganese ions. The mutant strain immobilized in calcium alginate beads was used in combination with surface-stabilized cultures for about 36-days in a continuous flow horizontal fermenter without any apparent loss in citric acid productivity. These results indicate that the manganese-resistant mutant is stable and may be used in the presence of sufficient manganese ions concentration (1.5 ppm) in the fermentation medium. This capability of the mutant strainA. niger GS-III has been correlated with greatly reduced levels (about one-thirds) of the NADP⁺ -isocitric dehydrogenase, one of the control points for citric acid accumulation.