Genome-wide screening of the genes required for tolerance to vanillin, which is a potential inhibitor of bioethanol fermentation, in Saccharomyces cerevisiae

<p>Abstract</p> <p>Background</p> <p>Lignocellulosic materials are abundant and among the most important potential sources for bioethanol production. Although the pretreatment of lignocellulose is necessary for efficient saccharification and fermentation, numerous by-products, including furan derivatives, weak acids, and phenolic compounds, are generated in the pretreatment step. Many of these components inhibit the growth and fermentation of yeast. In particular, vanillin is one of the most effective inhibitors in lignocellulose hydrolysates because it inhibits fermentation at very low concentrations. To identify the genes required for tolerance to vanillin, we screened a set of diploid yeast deletion mutants, which are powerful tools for clarifying the function of particular genes.</p> <p>Results</p> <p>Seventy-six deletion mutants were identified as vanillin-sensitive mutants. The numerous deleted genes in the vanillin-sensitive mutants were classified under the functional categories for 'chromatin remodeling' and 'vesicle transport', suggesting that these functions are important for vanillin tolerance. The cross-sensitivity of the vanillin-sensitive mutants to furan derivatives, weak acids, and phenolic compounds was also examined. Genes for ergosterol biosynthesis were required for tolerance to all inhibitory compounds tested, suggesting that ergosterol is a key component of tolerance to various inhibitors.</p> <p>Conclusion</p> <p>Our analysis predicts that vanillin tolerance in <it>Saccharomyces cerevisiae </it>is affected by various complicated processes that take place on both the molecular and the cellular level. In addition, the ergosterol biosynthetic process is important for achieving a tolerance to various inhibitors. Our findings provide a biotechnological basis for the molecular engineering as well as for screening of more robust yeast strains that may potentially be useful in bioethanol fermentation.</p

Initial Experience in Rectal Cancer Surgery for the Next Generation of Robotic Surgeons Trained in a Dual Console System

[Background]?Robotic surgery for rectal cancer is used worldwide, with an increasing incidence of robotic surgeons. Therefore, the most appropriate educational system for next-generation robotic surgeons should be urgently established. [Methods]?We analyzed 39 patients who underwent robotic rectal surgery performed by a next-generation surgeon with limited experienced in laparoscopic rectal cancer surgery. The dual console system was used in the initial 15 cases, and we assessed short-term outcomes and the learning curve on operative time using the cumulative sum method. [Results]?The patients were divided into two groups: 15 cases in the early phase, and 24 cases in the late phase. The operative time and surgeon console time were significantly shorter in the late phase than the early phase (P?< 0.001). Postoperative complications were more frequently observed in the early phase (P?= 0.049); however, the estimated blood loss and length of hospital stay were not significantly different. In the initial 15 cases that using the dual console, the average operative time changing to the expert surgeon was 82 minutes in the first 5 cases, 19 minutes on average in the next 5 cases, and no change occurred in the last 5 cases. The learning curve peaked after 14 cases, plateaued from case number 15 to 23, and decreased in a linear fashion until the final case. [Conclusion]?Education of a next generation surgeon using a dual console system for robotic rectal cancer surgery was performed safely

Kinetic analysis of the reaction catalyzed by chitinase A1 from Bacillus circulans WL-12 toward the novel substrates, partially N-deacetylated 4-methylumbelliferyl chitobiosides

AbstractThe kinetic behavior of chitinase A1 from Bacillus circulans WL-12 was investigated using the novel fluorogenic substrates, N-deacetylated 4-methylumbelliferyl chitobiosides [GlcN-GlcNAc-UMB (2), GlcNAc-GlcN-UMB (3), and (GlcN)2-UMB (4)], and the results were compared with those obtained using 4-methylumbelliferyl N,N′-diacetylchitobiose [(GlcNAc)2-UMB (1)] as the substrate. The chitinase did not release the UMB moiety from compound 4, but successfully released UMB from the other substrates. kcat/Km values determined from the releasing rate of the UMB moiety were: 145.3 for 1, 8.3 for 2, and 0.1 s−1 M−1 for 3. The lack of an N-acetyl group at subsite (−1) reduced the activity to a level 0.1% of that obtained with compound 1, while the absence of the N-acetyl group at subsite (−2) reduced the relative activity to 5.7%. These observations strongly support the theory that chitinase A1 catalysis occurs via a ‘substrate-assisted’ mechanism. Using these novel fluorogenic substrates, we were able to quantitatively evaluate the recognition specificity of subsite (−2) toward the N-acetyl group of the substrate sugar residue. The (−2) subsite of chitinase A1 was found to specifically recognize an N-acetylated sugar residue, but this specificity was not as strict as that found in subsite (−1)

Ruptured Small Intestinal Stromal Tumor Causing Concurrent Gastrointestinal and Intra-Abdominal Hemorrhage: A Case Report

Gastrointestinal stromal tumors (GISTs) originate from mesenchymal cells throughout the gastrointestinal tract. A common symptom is gastrointestinal hemorrhage; intra-abdominal hemorrhage is relatively rare. There are few reports of GIST presenting with both types of hemorrhage concurrently. A 77-year-old man was admitted to our hospital because of melena and anemia (Hb: 4.7 g/dL). Computed tomography revealed a small bowel tumor and high-density fluid in both the small intestine and the pelvic floor. We diagnosed a small intestinal tumor with concurrent gastrointestinal and intra-abdominal hemorrhage, and performed emergency surgery. The tumor arose from the small intestine and was ruptured. We found hemorrhage in the pelvic cavity and performed partial small intestine resection. Pathological findings revealed that the tumor was positive for c-Kit protein and was diagnosed as GIST. The patient was discharged from the hospital on postoperative day 9 and received imatinib 1 month postoperatively. We experienced a very rare case of ruptured GIST originating from the small intestine associated with both gastrointestinal and intra-abdominal hemorrhage. We also reviewed the relevant literature

Subsite structure of the endo-type chitin deacetylase from a deuteromycete, Colletotrichum lindemuthianum: an investigation using steady-state kinetic analysis and MS.

The endo-type chitin deacetylase (EC 3.5.1.41) from a deuteromycete, Colletotrichum lindemuthianum (ATCC 56676), catalyses the hydrolysis of the acetamido group of GlcNAc (2-acetamido-2-deoxy-D-glucose) residues in chitin or chito-oligosaccharides with a degree of polymerization (n) equal to or greater than 2. The steady-state kinetic parameters for the initial deacetylation reactions of (GlcNAc)(2-6) were determined using a direct, continuous spectrophotometric assay in combination with ESI-MS (electrospray ionization MS) analysis of the products. The dependence of the observed K(m) and k(cat)/K(m) on n suggests the presence of four enzyme subsites (-2, -1, 0 and +1) that interact with GlcNAc residues from the non-reducing end to the reducing end of the substrate. The turnover number (k (cat), 7 s(-1)) is independent of n and represents the intrinsic rate constant (k(int)) for the hydrolysis of the acetamido group in subsite 0. The subsite affinities for the GlcNAc residues were calculated from the observed k(cat)/K(m) values (A (-2), -11.0; A (-1), -1.5; A (0), -7.7; A (+1), -12.5 kJ x mol(-1)). The increments in the subsite affinities due to the recognition of the acetamido groups were calculated [DeltaDelta G ((N-acetyl))=3.3, 0, 4.0 and 0 kJ x mol(-1) for subsites -2, -1, 0 and +1 respectively]. The steady-state kinetic parameters for the second deacetylation reaction of (GlcNAc)(4) were also determined using (GlcNAcGlcNAcGlcNGlcNAc) as the substrate. The comparison of the experimental and theoretical values (calculated using the subsite affinities) suggests that the mono-deacetylated substrate binds strongly in a non-productive mode occupying all four subsites, thereby inhibiting the second deacetylation reaction

Comparison of textural properties and structure of gels prepared from cooked rice grain under different conditions

The objective of this study was to investigate the effects of rice variety, water content, and preparation temperature on the textural properties of gels processed from cooked rice grains via high‐speed shear homogenization. Rice gels were prepared from seven high‐amylose rice varieties. The results demonstrated the significant differences in rice gel hardness and hardening rates during storage based on the rice variety used. The proportion of short chains of amylopectin was negatively correlated with the hardness of the rice gel. The sample temperature before shear treatment also influenced the rice gel hardness. Rice gels prepared from cooked rice maintained at 75°C prior to homogenization showed a higher breaking force than those from cooked rice at 25°C. Observation using scanning electron microscopy demonstrated the tendency of the cooked rice sample maintained at 75°C to form a finer gel network after homogenization than those at 25°C from the same rice variety