Acid-catalyzed steam pretreatment of lodgepole pine and subsequent enzymatic hydrolysis and fermentation to ethanol

Utilization of ethanol produced from biomass has the potential to offset the use of gasoline and reduce CO(2) emissions. This could reduce the effects of global warming, one of which is the current outbreak of epidemic proportions of the mountain pine beetle (MPB) in British Columbia (BC), Canada. The result of this is increasing volumes of dead lodgepole pine with increasingly limited commercial uses. Bioconversion of lodgepole pine to ethanol using SO(2)-catalyzed steam explosion was investigated. The optimum pretreatment condition for this feedstock was determined to be 200 degrees C, 5 min, and 4% SO(2) (w/w). Simultaneous saccharification and fermentation (SSF) of this material provided an overall ethanol yield of 77% of the theoretical yield from raw material based on starting glucan, mannan, and galactan, which corresponds to 244 g ethanol/kg raw material within 30 h. Three conditions representing low (L), medium (M), and high (H) severity were also applied to healthy lodgepole pine. Although the M severity conditions of 200 degrees C, 5 min, and 4% SO(2) were sufficiently robust to pretreat healthy wood, the substrate produced from beetle-killed (BK) wood provided consistently higher ethanol yields after SSF than the other substrates tested. BK lodgepole pine appears to be an excellent candidate for efficient and productive bioconversion to ethanol.     

On the synthesis of the 2,6-dideoxysugar L-digitoxose

As deoxysugars are integral components of many natural products, the development of efficient chemical and enzymatic routes to prepare these compounds is of particular interest. Herein, we report a comparison of several synthetic methodologies used to prepare protected derivatives of the 2,6-dideoxysugar l-digitoxose. A novel, stereoselective synthetic route to efficiently access methyl 4-O-tert-butyldimethylsilyl-2,6-dideoxy-3-O-trimethylsilyl-alpha-l-ribo-hexopyranoside in 35% yield over nine facile steps is described.     

Autonomy of cell proliferation and developmental programs during Arabidopsis aboveground organ morphogenesis

Elaboration of size and shape in multicellular organisms involves coordinated cell division and cell growth. In higher plants, continuity of cell layer structures exists from the shoot apical meristem (SAM), where organ primordia arise, to mature aboveground organs. To unravel the extent of inter-cell layer coordination during SAM and aboveground organ development, cell division in the epidermis was selectively restricted by expressing two cyclin-dependent kinase inhibitor genes, KRP1/ICK1 and KRP4, driven by the L1 layer-specific AtML1 promoter. The transgenes conferred reduced plant size with striking, distorted lateral organ shape. While epidermal cell division was severely inhibited with compensatory cell size enlargement, the underlying mesophyll/cortex layer kept normal cell numbers and resulted in small, packed cells with disrupted cell files. Our results demonstrate the autonomy of cell number checkpoint in the underlying tissues when epidermal cell division is restricted. Finally, the L1 layer-specific expression of both KRP1/ICK1 and KRP4 showed no effects on the structure and function of the SAM, suggesting that the effects of these cyclin-dependent kinase inhibitors are context dependent.     

Specialized mismatch repair function of Glu339 in the Phe-X-Glu motif of yeast Msh6

The major eukaryotic mismatch repair (MMR) pathway requires Msh2-Msh6, which, like Escherichia coli MutS, binds to and participates in repair of the two most common replication errors, single base-base and single base insertion-deletion mismatches. For both types of mismatches, the side chain of E. coli Glu38 in a conserved Phe-X-Glu motif interacts with a mismatched base. The Ovarepsilon of Glu38 forms a hydrogen bond with either the N7 of purines or the N3 of pyrimidines. We show here that changing E. coli Glu38 to alanine results in nearly complete loss of repair of both single base-base and single base deletion mismatches. In contrast, a yeast strain with alanine replacing homologous Glu339 in Msh6 has nearly normal repair for insertion-deletion and most base-base mismatches, but is defective in repairing base-base mismatches characteristic of oxidative stress, e.g. 8-oxo-G.A mismatches. The results suggest that bacterial MutS and yeast Msh2-Msh6 differ in how they recognize and/or process replication errors involving undamaged bases, and that Glu339 in Msh6 may have a specialized role in repairing mismatches containing oxidized bases.     

Consensus framework for exploring microarray data using multiple clustering methods

The large variety of clustering algorithms and their variants can be daunting to researchers wishing to explore patterns within their microarray datasets. Furthermore, each clustering method has distinct biases in finding patterns within the data, and clusterings may not be reproducible across different algorithms. A consensus approach utilizing multiple algorithms can show where the various methods agree and expose robust patterns within the data. In this paper, we present a software package - Consense, written for R/Bioconductor - that utilizes such an approach to explore microarray datasets. Consense produces clustering results for each of the clustering methods and produces a report of metrics comparing the individual clusterings. A feature of Consense is identification of genes that cluster consistently with an index gene across methods. Utilizing simulated microarray data, sensitivity of the metrics to the biases of the different clustering algorithms is explored. The framework is easily extensible, allowing this tool to be used by other functional genomic data types, as well as other high-throughput OMICS data types generated from metabolomic and proteomic experiments. It also provides a flexible environment to benchmark new clustering algorithms. Consense is currently available as an installable R/Bioconductor package (http://www.ohsucancer.com/isrdev/consense/).     

Asymmetric deceleration of ClpB or Hsp104 ATPase activity unleashes protein-remodeling activity

Two members of the AAA+ superfamily, ClpB and Hsp104, collaborate with Hsp70 and Hsp40 to rescue aggregated proteins. However, the mechanisms that elicit and underlie their protein-remodeling activities remain unclear. We report that for both Hsp104 and ClpB, mixtures of ATP and ATP-gammaS unexpectedly unleash activation, disaggregation and unfolding activities independent of cochaperones. Mutations reveal how remodeling activities are elicited by impaired hydrolysis at individual nucleotide-binding domains. However, for some substrates, mixtures of ATP and ATP-gammaS abolish remodeling, whereas for others, ATP binding without hydrolysis is sufficient. Remodeling of different substrates necessitates a diverse balance of polypeptide 'holding' (which requires ATP binding but not hydrolysis) and unfolding (which requires ATP hydrolysis). We suggest that this versatility in reaction mechanism enables ClpB and Hsp104 to reactivate the entire aggregated proteome after stress and enables Hsp104 to control prion inheritance.