Heterologous expression and biochemical characterization of a thermostable xylulose kinase from Bacillus coagulans IPE22.

Xylulose kinase is an important enzyme involved in xylose metabolism, which is considered as essential biocatalyst for sustainable lignocellulosic-derived pentose utilization. Bacillus coagulans IPE22 is an ideal bacterium for refinery due to its strong ability to ferment xylose at high temperature. However, the B. coagulans xylose utilization mechanism remains unclear and the related promising enzymes need to be developed. In the present study, the gene coding for xylulose kinase from B. coagulans IPE22 (Bc-XK) was expressed in Escherichia coli BL21 (DE3). Bc-XK has a 1536 bp open reading frame, encoding a protein of 511 amino acids (56.15 kDa). Multiple sequence alignments were performed and a phylogenetic tree was built to evaluate differences among Bc-XK and other bacteria homologs. Bc-XK showed a broad adaptability to high temperature and the enzyme displayed its best performance at pH 8.0 and 60 °C. Bc-XK was activated by Mg2+ , Mn2+ , and Co2+ . Meanwhile, the enzyme could keep activity at 60 °C for at least 180 min. KM values of Bc-XK for xylulose and ATP were 1.29 mM and 0.76 mM, respectively. The high temperature stability of Bc-XK implied that it was an attractive candidate for industrial application.

Expression of inulinase gene in the oleaginous yeast Yarrowia lipolytica and single cell oil production from inulin-containing materials.

Yarrowia lipolytica ACA-DC 50109 has been reported to be an oleaginous yeast and significant quantities of lipids were accumulated inside the yeast cells. In this study, the INU1 gene encoding exo-inulinase cloned from Kluyveromyces marxianus CBS 6556 was ligated into the expression plasmid pINA1317 and expressed in the cells of the oleaginous yeast. The activity of the inulinase with 6 × His tag secreted by the transformant Z31 obtained was found to be 41.7U mL(-1) after cell growth for 78 h. After optimization of the medium and cultivation conditions for single cell oil production, the transformant could accumulate 46.3% (w/w) oil from inulin in its cells and cell dry weight was 11.6 g L(-1) within 78 h at the flask level. During the 2-L fermentation, the transformant could accumulate 48.3% (w/w) oil from inulin in its cells and cell dry weight was 13.3 g L(-1) within 78 h while the transformant could accumulate 50.6% (w/w) oil from extract of Jerusalem artichoke tubers in its cells and cell dry weight was 14.6 g L(-1) within 78 h. At the end of fermentation, most of the added sugar was utilized by the transformant cells. Over 91.5% of the fatty acids from the transformant cultivated in the extract of Jerusalem artichoke tubercles was C(16:0), C(18:1) and C(18:2), especially C(18:1) (58.5%).

Single cell protein production from yacon extract using a highly thermosensitive and permeable mutant of the marine yeast Cryptococcus aureus G7a and its nutritive analysis.

The intracellular protein in the highly thermosensitive and permeable mutant can be easily released when they are incubated both in the low-osmolarity water and at the non-permissive temperature (usually 37 degrees C). After the mutant was grown in the yacon extract for 45 h, the crude protein content in the highly thermosensitive and permeable mutant Z114 was 59.1% and over 61% of the total protein could be released from the cells treated at 37 degrees C. The mutant cells grown in the yacon extract still contained high level of essential amino acids and other nutrients. This means that the yacon extract could be used as the medium for growth of the highly thermosensitive and permeable mutant which contained high content of crude protein.

Taxonomic importance of pollen morphology in Veratrum L. (Melanthiaceae) using microscopic techniques.

Previous studies reported that intrageneric relationships of genus Veratrum of family Melanthiaceae are controversial and hard to delimit. Therefore, we observed the pollen morphological features of six species in the genus Veratrum in detail using both light microscopy and scanning electron microscopy and investigated their significance for Veratrum taxonomy. Among them, five were studied for the first time. The results demonstrated that pollen grains of Veratrum are medium in size with P/E being 0.31-0.60. Three types of shape in polar view have been observed elliptic, long-elliptic, or wide-elliptic with blunt or rounded at both ends. Two types of width of colpus reported narrow or wide, two types of depth of colpus reported deep or flat, and two types of length of colpus reported extend almost or do not extend to the ends, whereas two types of colpus membranes reported absent or obvious. One type of surface ornamentation was noted as reticulate. These results support species Veratrum album and Veratrum lobelianum as well as Veratrum grandiflorum and Veratrum oxysepalum as two independent species, respectively, rather than classifying Veratrum into two sections. Overall, we demonstrated that the ratio of polar axis length to equatorial axial length, pollen characteristics at the polar view, the colpus morphology, and the surface ornamentation of pollen grains of genus Veratrum have important systematic significance in identification and delimitation of species.

Pollen morphology of Aletris L. (Nartheciaceae) and its systematic significance.

In this paper, we studied pollen morphologies of seven species in genus Aletris in detail by light microscopy and scanning electron microscopy. Of these, six species were reported for the first time. The palynological characteristics do not support the infrageneric classification into two clades. The results indicated that pollen grains of Aletris are small or medium with the P/E ratio of 0.36-0.59. They are elliptic or long-elliptic in the polar view with blunt, round or acute ends and bilateral symmetric with a monosulcate, narrow or wide, deep colpus that has length extending to the ends of pollen grains, obvious or absent colpus membranes. The pollen ornamentation is gemmate, perforated, or reticulate. The sexine is slightly or quite thicker than the nexine.

Cellulose nanocrystals derived from Enteromorpha prolifera and their use in developing bionanocomposite films with water-soluble polysaccharides extracted from E. prolifera.

The biomass from Enteromorpha prolifera is used to exploit and transfer this common green bloom algae into new value-added products. In this work, cellulose nanocrystals (CNCs) were first prepared with the cellulose extracted from the biomass of E. prolifera. Initially, cellulose was treated by alkali and bleaching treatments. Subsequently, CNCs were isolated by acid hydrolysis with different concentrations. Structural changes and crystallinity were characterized by Fourier transform infrared spectroscopy (FTIR) spectroscopy, X-ray diffraction (XRD) analysis and t thermogravimetric analysis. The crystallinity index and crystallite size of the prepared CNCs depended mainly on acid concentrations. TEM observation revealed that the CNCs obtained by hydrolysis with 60% acid (CNC60) exhibited a 'needle-like' nanocrystals with a length of 177 ±â€¯12 nm and width of 3 ±â€¯1 nm, indicating that the CNC60 had a high ratio aspect and could serve as reinforcement nanofiller. Further investigation showcased that the addition of CNC60 exhibited the best effect for enhancing the mechanical properties of bionanocomposite films derived from the water-soluble polysaccharides extracted from the biomass of E. prolifera. Therefore, a new strategy was offered in this study for the comprehensive utilization of E. prolifera biomass to fabricate them into high-strength film materials.

Biotechnological potential of inulin for bioprocesses.

Inulin consists of linear chains of ß-2,1-linked D-fructofuranose molecules terminated by a glucose residue through a sucrose-type linkage at the reducing end. In this review article, inulin and its applications in bioprocesses are overviewed. The tubers of many plants, such as Jerusalem artichoke, chicory, dahlia, and yacon contain a large amount of inulin. Inulin can be actively hydrolyzed by microbial inulinases to produce fructose, glucose and inulooligosaccharides (IOS). The fructose and glucose formed can be further transformed into ethanol, single-cell protein, single cell oil and other useful products by different microorganisms. IOS formed have many functions. Therefore, inulin can be widely used in food, feed, pharmaceutical, chemical and biofuels industries.

Direct conversion of inulin and extract of tubers of Jerusalem artichoke into single cell oil by co-cultures of Rhodotorula mucilaginosa TJY15a and immobilized inulinase-producing yeast cells.

In this study, it was found that the immobilized inulinase-producing cells of Pichia guilliermondii M-30 could produce 169.3 U/ml of inulinase activity while the free cells of the same yeast strain only produced 124.3 U/ml of inulinase activity within 48 h. When the immobilized inulinase-producing yeast cells were co-cultivated with the free cells of Rhodotorula mucilaginosa TJY15a, R. mucilaginosa TJY15a could accumulate 53.2% oil from inulin in its cells and cell dry weight reached 12.2g/l. Under the similar conditions, R. mucilaginosa TJY15a could accumulate 55.4% (w/w) oil from the extract of Jerusalem artichoke tubers in its cells and cell dry weight reached 12.8 g/l within 48 h. When the co-cultures were grown in 2l fermentor, R. mucilaginosa TJY15a could accumulate 56.6% (w/w) oil from the extract of Jerusalem artichoke tubers in its cells and cell dry weight reached 19.6g/l within 48 h. Over 90.0% of the fatty acids from the yeast strain TJY15a grown in the extract of Jerusalem artichoke tubers was C(16:0), C(18:1) and C(18:2), especially C(18:1) (50.6%).