Correlated rates of synonymous site evolution across plant genomes

Synonymous substitution rates have been shown to vary among evolutionary lineages of both nuclear and organellar genes across a broad range of taxonomic groups. In animals, rate heterogeneity does not appear to be correlated across nuclear and mitochondrial genes. In this paper, we contrast substitution rates in two plant groups and show that grasses evolve more rapidly than palms at synonymous sites in a mitochondrial, a nuclear, and a plastid gene. Furthermore, we show that the relative rates of synonymous substitution between grasses and palms are similar at the three loci. The correlation in synonymous substitution rates across genes is particularly striking because the three genes evolve at very different absolute rates. In contrast, relative rates of nonsynonymous substitution are not conserved among the three genes.      

Conversion of corn milling fibrous co-products into ethanol by recombinant Escherichia coli strains K011 and SL40

Corn hulls and corn germ meal were both evaluated as feedstocks for production of ethanol for biofuel. Currently, these fibrous co-products are combined with corn steep liquor and the fermentation bottoms (if available) and marketed as cattle feed. Samples were obtained from wet and dry corn mills. The corn hulls and germ meal were evaluated for starch and hemicellulose compositions. Starch contents were 12 to 32% w/w and hemicellulose (arabinoxylans) contents were 23 to 64% w/w. Corn fibrous samples were hydrolysed, using dilute sulphuric acid, into mixed sugar streams containing arabinose, glucose and xylose. Total sugar concentrations in the hydrolysate varied from 8.4 to 10.8% w/v. The hydrolysates were fermented to ethanol using recombinant E. coli strains K011 and SL40. Ethanol yields were 0.38 to 0.41g ethanol produced/g total sugars consumed and fermentations were completed in 60h or less. However, residual xylose was detected for each hydrolysate fermentation and was especially significant for fermentations using strain SL40. Strain K011 was a superior ethanologenic strain compared with strain SL40 in terms of both ethanol yield and maximum productivity.     

Engineering polyphosphate metabolism in Escherichia coli: implications for bioremediation of inorganic contaminants

Polyphosphate metabolism plays an important role in the bioremediation of phosphate contamination in municipal wastewater, and may play a key role in heavy metal tolerance and bioremediation. However, little is known about the regulation of polyphosphate metabolism in microorganisms and its role in heavy metal toxicity. We have manipulated polyphosphate metabolism in Escherichia coli by overexpressing the genes for polyphosphate kinase (ppk) and for polyphosphatase (ppx) under control of their native promoters and inducible promoters. Overexpression of ppk results in high levels of intracellular polyphosphate, improved phosphate uptake, but no increase in tolerance to heavy metals. Overexpression of both ppk and ppx results in lower levels of intracellular polyphosphate, secretion of phosphate from the cell, and increased tolerance to heavy metals. Metabolic flux analysis indicates that the cell responds to increased flux through the PPK-PPX pathway by altering flux through the TCA cycle.     

The role of site-specific N-glycosylation in secretion of soluble forms of rabies virus glycoprotein

Rabies virus glycoprotein is important in the biology and pathogenesis of neurotropic rabies virus infection. This transmembrane glycoprotein is the only viral protein on the surface of virus particles, is the viral attachment protein that facilitates virus uptake by the infected cell, and is the target of the host humoral immune response to infection. The extracellular domain of this glycoprotein has N- glycosylation sequons at Asn37, Asn247, and Asn319. Appropriate glycosylation of these sequons is important in the expression of the glycoprotein. Soluble forms of rabies virus glycoprotein were constructed by insertion of a stop codon just external to the transmembrane domain. Using site-directed mutagenesis and expression in transfected eukaryotic cells, it was possible to compare the effects of site-specific glycosylation on the cell-surface expression and secretion of transmembrane and soluble forms, respectively, of the same glycoprotein. These studies yielded the surprising finding that although any of the three sequons permitted cell surface expression of full-length rabies virus glycoprotein, only the N-glycan at Asn319 permitted secretion of soluble rabies virus glycoprotein. Despite its biological and medical importance, it has not yet been possible to determine the crystal structure of the full-length transmembrane form of rabies virus glycoprotein which contains heterogeneous oligosaccharides. The current studies demonstrate that a soluble form of rabies virus glycoprotein containing only one sequon at Asn319 is efficiently secreted in the presence of the N-glycan processing inhibitor 1-deoxymannojirimycin. Thus, it is possible to purify a conformationally relevant form of rabies virus glycoprotein that contains only one N-glycan with a substantial reduction in its microheterogeneity. This form of the glycoprotein may be particularly useful for future studies aimed at elucidating the three-dimensional structure of this important glycoprotein.      

Ethanol production from AFEX pretreated corn fiber by recombinant bacteria

Summary Fermentation of an enzymatic hydrolyzate of ammonia fiber explosion (AFEX) pretreated corn fiber (containing a mixture of different sugars including glucose, xylose, arabinose, and galactose) by genetically-engineered Escherichia coli strain SL40 and KO11 and Klebsiella oxytoca strain P2 was investigated under pH-controlled conditions. Both E. coli strains (SL40 and KO11) efficiently utilized most of the sugars contained in the hydrolyzate and produced a maximum of 26.6 and 27.1 g/l ethanol, respectively, equivalent to 90 and 92% of the theoretical yield. Very little difference was observed in cell growth and ethanol production between fermentations of the enzymatic hydrolyzate and mixtures of pure sugars, simulating the hydrolyzate. These results confirm the fermentability of the AFEX-treated corn fiber hydrolyzate by ethanologenic E. coli. K.oxytoca strain P2, on the other hand, showed comparatively poor growth and ethanol production (maximum 20 g/l) from both enzymatic hydrolyzate and simulated sugar mixtures under the same fermentation conditions.     

A survey of yeast from the Yarrowia clade for lipid production in dilute acid pretreated lignocellulosic biomass hydrolysate

Applied Microbiology and Biotechnology - Yarrowia lipolytica is an oleaginous yeast species that has attracted attention as a model organism for synthesis of single cell oil. Among over 50 isolates...     

Effect of polyphosphate metabolism on the Escherichia coli phosphate-starvation response.

A previously developed dynamic model of the Escherichia coli Pho regulon was extended to investigate the effect of polyphosphate synthesis and degradation on this control system. Differential equations for ATP and polyphosphate were formulated, and the model was applied to the growth of cells containing the ppk and ppx genes under control of separate, inducible promoters. In agreement with recent experimental observations, the degradation of polyphosphate by PPX during a period of phosphate limitation could repress the phosphate-starvation response. This is attributed to the release of phosphate from the cell into the periplasm, where it can be detected by the external phosphate sensor. A segregated model was then developed to account for differences in K(I), the dissociation constant for the repression complex, among cells of the population. Since K(I) is the key parameter in determining whether the Pho response is induced or repressed at a particular surface phosphate concentration, this permitted the induction of some cells while others remained repressed. The induction profiles resulting from the population-averaged values more closely matched experimental results than did those with the nonsegregated model.