Yinbo Qu

Enzyme-modified nanoporous gold-based electrochemical biosensors

On the basis of the unique physical and chemical properties of nanoporous gold (NPG), which was obtained simply by dealloying Ag from Au/Ag alloy, an attempt was made in the present study to develop NPG-based electrochemical biosensors. The NPG-modified glassy carbon electrode (NPG/GCE) exhibited high-electrocatalytic activity toward the oxidation of nicotinamide adenine dinucleotide (NADH) and hydrogen peroxide (H(2)O(2)), which resulted in a remarkable decrease in the overpotential of NADH and H(2)O(2) electro-oxidation when compared with the gold sheet electrode. The high density of edge-plane-like defective sites and large specific surface area of NPG should be responsible for the electrocatalytic behavior. Such electrocatalytic behavior of the NPG/GCE permitted effective low-potential amperometric biosensing of ethanol or glucose via the incorporation of alcohol dehydrogenase (ADH) or glucose oxidase (GOD) within the three-dimensional matrix of NPG. The ADH- and GOD-modified NPG-based biosensors showed good analytical performance for biosensing ethanol and glucose due to the clean, reproducible and uniformly distributed microstructure of NPG. The stabilization effect of NPG on the incorporated enzymes also made the constructed biosensors very stable. After 1 month storage at 4 degrees C, the ADH- and GOD-based biosensors lost only 5.0% and 4.2% of the original current response. All these indicated that NPG was a promising electrode material for biosensors construction.

High concentration ethanol production from corncob residues by fed-batch strategy

Ethanol production from corncob residues (CCR) pretreated by different methods was studied. The structure features of these CCR were analyzed by Fourier transform-infrared spectrum (FT-IR), X-ray diffraction (XRD), and field emission scanning electron microscope (SEM). Simultaneous saccharification and fermentation (SSF) was performed by adding crude cellulase preparations from Penicillium decumbens JUA10-1 at 30 degrees C. The results suggested that different pretreatments resulted in different composition and structure of residues; these changes had a significant influence on ethanol productivity and concentration. The fed-batch method was combined with SSF to enhance ethanol concentration further and reduce enzyme dosage. Moreover, the absorption and desorption phenomena of cellobiohydrolase I (CBH I) (70 kDa) were observed to be related to lignin contents in residues. These results demonstrated that despite the application of low enzyme dosage, high concentration ethanol could be produced from pretreated corncobs by combining fed-batch method with SSF.

Genomic and Secretomic Analyses Reveal Unique Features of the Lignocellulolytic Enzyme System of Penicillium decumbens

Many Penicillium species could produce extracellular enzyme systems with good lignocellulose hydrolysis performance. However, these species and their enzyme systems are still poorly understood and explored due to the lacking of genetic information. Here, we present the genomic and secretomic analyses of Penicillium decumbens that has been used in industrial production of lignocellulolytic enzymes in China for more than fifteen years. Comparative genomics analysis with the phylogenetically most similar species Penicillium chrysogenum revealed that P. decumbens has evolved with more genes involved in plant cell wall degradation, but fewer genes in cellular metabolism and regulation. Compared with the widely used cellulase producer Trichoderma reesei, P. decumbens has a lignocellulolytic enzyme system with more diverse components, particularly for cellulose binding domain-containing proteins and hemicellulases. Further, proteomic analysis of secretomes revealed that P. decumbens produced significantly more lignocellulolytic enzymes in the medium with cellulose-wheat bran as the carbon source than with glucose. The results expand our knowledge on the genetic information of lignocellulolytic enzyme systems in Penicillium species, and will facilitate rational strain improvement for the production of highly efficient enzyme systems used in lignocellulose utilization from Penicillium species.

Status and prospect of lignocellulosic bioethanol production in China

As a developing country with the largest population, China faces a serious challenge in satisfying its continuously increasing energy demand, especially for liquid fuel. Bioethanol production from lignocellulosic material is a potential and feasible method to solve the many problems in China, and it was supported by the Chinese government. Many research projects in China on lignocellulosics ethanol production have been carried out. After more than 30years of research, several pilot scale facilities have been constructed. This review focuses on the recent research activities and developments in lignocellulosic ethanol production during the past decade in China. As case study, a corncob biorefinery process is introduced.

The Effects of Wheat Bran Composition on the Production of Biomass-Hydrolyzing Enzymes by Penicillium decumbens

The effects of the starch, protein, and soluble oligosaccharides contents in wheat bran on the extracellular biomass-hydrolyzing enzymes activities released by Penicillium decumbens mycelia grown in batch fermentations have been examined. The results showed increased starch content correlated directly with an increase in released amylase activity but inversely with the levels of secreted cellulase and xylanase. High amounts of protein in wheat bran also reduced the activities of cellulase, xylanase and protease in the culture medium. The effects of the soluble and insoluble components of wheat bran and cello-oligosaccharides supplements on production of extracellular cellulase and xylanase were compared. The soluble cello-oligosaccharides compositions in wheat bran were proved to be one of the most significant factors for cellulase production. According to the results of this research, determining and regulating the composition of wheat bran used as a fermentation supplement may allow for improved induction of cellulase and xylanase production.

Sophorolipid produced from the new yeast strain Wickerhamiella domercqiae induces apoptosis in H7402 human liver cancer cells

The effects of sophorolipid on the growth and apoptosis of H7402 human liver cancer cells were investigated. By treatment with sophorolipid, a dose- and time-dependent inhibition of cell proliferation was observed. The cells developed many features of apoptosis, including condensation of chromatin, nuclear fragmentation, and appearance of apoptotic bodies, and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling positive cells were stained dark brown. Sophorolipid treatment induced apoptosis in H7402 cells by blocking cell cycle at G1 phase and partly at S phase, activating caspase-3, and increasing Ca2+ concentration in cytoplasm. These findings may suggest a potential use of sophorolipid for liver cancer treatment.

Differences in the adsorption of enzymes onto lignins from diverse types of lignocellulosic biomass and the underlying mechanism

BackgroundNon-productive cellulase adsorption onto lignin has always been deemed to negatively affect the enzymatic hydrolysis of lignocellulosic feedstocks. Therefore, understanding enzyme-lignin interactions is essential for the development of enzyme mixtures, the processes of lignocellulose hydrolysis, and the genetic modification of lignocellulosic biomass and enzymes. In this work, we examined the properties of six lignins from diverse types of lignocellulosic biomass (aspen, pine, corn stover, kenaf, and two Arabidopsis lines, wild-type and SALK mutant of fah1) to determine the mechanism of differences in their adsorption of enzymes.ResultsWe found that lignin sources affected enzyme adsorption using structural features, such as functional groups and lignin composition. Guaiacyl (G) lignin had a higher adsorption capacity on enzymes than syringyl (S) lignin. The low S/G ratio and high uniform lignin fragment size had good correlations with high adsorption capacity. A higher content of phenolic hydroxyl groups and a lower content of carboxylic acid groups resulted in stronger adsorption affinity for corn stover lignin (CL) than for kenaf lignin (KL) and aspen lignin (AL). The lower amount of aliphatic hydroxyls that reduced hydrophobic interactions could explain the higher adsorption capacity of pine lignin (PL) than CL. Enzyme activity assays, as well as the hydrolysis of Avicel, phosphoric acid-swollen cellulose (PASC), and holocellulose, were performed to study the behaviors of mono-component enzymes that resulted in adsorption. We found that cellobiohydrolase (CBH) and xylanase were adsorbed the most by all lignins, endoglucanase (EG) showed less inhibition, and β-glucosidase (BG) was the least affected by lignins, indicating the important role of carbohydrate-binding module (CBM) in protein adsorption.ConclusionLignin sources affect enzyme adsorption using structural features and lignin composition, such as S/G ratio, carboxylic acid, aliphatic hydroxyl, and phenolic hydroxyl. For mono-component enzymes, the adsorption capacity decreased in the order CBH, xylanase > EG > BG. These investigations revealed the difference in lignin properties between diverse biomass and adsorption capacity of enzymes to lignins, and the possible underlying mechanism. The results can also serve as a reference for the genetic modification of lignocellulosic biomass and enzymes.

Genome shuffling improves production of cellulase by Penicillium decumbens JU-A10

Aims:  Improvement of cellulase production of Penicillium decumbens by genome shuffling of an industrial catabolite‐repression‐resistant strain JU‐A10 with its mutants.

Engineering endoglucanase II from Trichoderma reesei to improve the catalytic efficiency at a higher pH optimum

The catalytic efficiency and pH optimum of Trichoderma reesei endo-beta-1,4-glucanase II were improved by protein engineering. We subjected residue 342 to saturation mutagenesis, and further changed the enzyme by random mutagenesis and two rounds of DNA shuffling. Enzyme variants were purified and characterized. Variant N342V exhibited an optimal activity at pH 5.8, corresponding to a basic shift of 1 pH unit compared with the wild-type enzyme, and had improved catalytic efficiency (1.5-fold of k(cat)/K(m)) for the main substrates at pH 6.2. Variants N342R and N39R/L218H/W276R/N342T both had a pH optimum of 6.2 and the latter had improved catalytic efficiency (1.4-fold of k(cat)/K(m)) at pH 6.2. Variants L218H, Q139R/N342T and Q139R/L218H/W276R/N342T all had more than 4.5-fold higher activity in reactions compared with the wild-type at pH 7.0. The relationship between the structures and the activities of the variants were analyzed by modeling the structures of the endoglucanase II variants. More stable helixes and changed electrostatic interactions between the catalytic residues and substrates may explain the higher activities and higher pH optima of the variants.

Isolation and characterization of a β-glucosidase from Penicillium decumbens and improving hydrolysis of corncob residue by using it as cellulase supplementation

A β-glucosidase from Penicillium decumbens was purified and characterized. The enzyme presented as a single band of 120kDa on SDS-PAGE, showed optimal temperature of 65-70°C and optimal pH of 4.5-5.0. The β-glucosidase showed relatively higher affinity to pNPG and the highest affinity to salicin with the Km value as 0.0064 and 0.0188mM, respectively. The gene coding for it was obtained with an ORF of 2586bp coding for 861 amino acids belonging to glycoside hydrolases family 3. The purified enzyme could improve the saccharifying ability of cellulose when it was added to the cellulase systems of Trichoderma reesei QM 9414. The several properties of it, including its pH and temperature optima, the high affinity to substrates and high specific activity, make it has great potential to be utilized as supplementation in conversion of corncob residue and other lignocellulosic biomass into simple sugars.