Expression of soluble moloney murine leukemia virus-reverse transcriptase in Escherichia coli BL21 star (DE3) using autoinduction system.

Autoinduction systems in Escherichia coli can control the production of proteins without the addition of a particular inducer. In the present study, we optimized the heterologous expression of Moloney Murine Leukemia Virus derived Reverse Transcriptase (MMLV-RT) in E. coli. Among 4 autoinduction media, media Imperial College resulted the highest MMLV-RT overexpression in E. coli BL21 Star (DE3) with incubation time 96 h. The enzyme was produced most optimum in soluble fraction of lysate cells. The MMLV-RT was then purified using the Immobilized Metal Affinity Chromatography method and had specific activity of 629.4 U/mg. The system resulted lower specific activity and longer incubation of the enzyme than a classical Isopropyl ß-D-1-thiogalactopyranoside (IPTG)-induction system. However, the autoinduction resulted higher yield of the enzyme than the conventional induction (27.8%). Techno Economic Analysis revealed that this method could produce MMLV-RT using autoinduction at half the cost of MMLV-RT production by IPTG-induction. Bioprocessing techniques are necessary to conduct to obtain higher quality of MMLV-RT under autoinduction system.

Construction of Novel Yeast Strains from Candida tropicalis KBKTI 10.5.1 and Saccharomyces cerevisiae DBY1 to Improve the Performance of Ethanol Production Using Lignocellulosic Hydrolysate.

Increased consumption of xylose-glucose and yeast tolerance to lignocellulosic hydrolysate are the keys to the success of second-generation bioethanol production. Candida tropicalis KBKTI 10.5.1 is a new isolated strain that has the ability to ferment xylose. In contrast to Saccharomyces cerevisiae DBY1 which only can produce ethanol from glucose fermentation. The research objective is the application of the genome shuffling method to increase the performance of ethanol production using lignocellulosic hydrolysate. Mutants were selected on xylose and glucose substrates separately and using random amplified polymorphic DNA (RAPD) analysis. The ethanol production using lignocellulosic hydrolysate by parents and mutants was evaluated using a batch fermentation system. Concentrations of ethanol, residual sugars, and by-products such as glycerol, lactate and acetate were measured using HPLC machine equipped with Hiplex H for carbohydrate column and a refraction index detector (RID). Ethanol produced by Fcs1 and Fcs4 mutants on acid hydrolysate increased by 26.58% and 24.17% from parent DBY1, by 14.94% and 21.84% from parent KBKTI 10.5.1. In contrast to the increase in ethanol production on alkaline hydrolysate, Fcs1 and Fcs4 mutants only experienced an increase in ethanol production by 1.35% from the parent KBKTI 10.5.1. Ethanol productivity by Fcs1 and Fcs4 mutants on acid hydrolysate reached 0.042 g/L/h and 0.044 g/L/h. The recombination of the genomes of different yeast species resulted in novel yeast strains that improved resistance performance and ethanol production on lignocellulosic hydrolysates.

Effect of Ethanol-Derived Clove Leaf Extract on the Oxidative Stress Response in Yeast Schizosaccharomyces pombe.

Compared to the widely explored antioxidant activity from the clove bud extract, less data are available regarding the potential pharmacological use of clove leaves. Our study aimed to explore the antioxidant activity of clove leaves extract in the cellular level. Thus, we used the yeast Schizosaccharomyces pombe as model organisms. Our data indicate that, following extract treatment (100 ppm), the viability of the stationary phase cells of S. pombe was higher than without extract and that of calorie restriction treatments. 100 ppm extract treatment also increased cell viability against H2O2-induced oxidative stress. Those data indicate that the extract could promote oxidative stress tolerance response in yeast cells, which occurred either during the stationary phase or due to exogenous exposure. Higher dose of extract (500 ppm) showed opposite effects, as cell viability was lower than that without treatment. Analysis toward the mitochondrial activity revealed that the extract did not induce mitochondrial activity unlike the calorie restriction treatment. Based on our data, clove leaf extract promotes oxidative stress tolerance response in the yeast S. pombe, independent to that mitochondrial adaptive ROS signaling which commonly occurs in calorie restriction-induced oxidative stress tolerance response.

Diversity of lactic acid bacterial in inasua fermentation.

BACKGROUND AND OBJECTIVES: Inasua is one of the traditional fermented fish products in Maluku, Indonesia. There are two types of inasua, i.e. with and without sap. The research aimed to study the succession of lactic acid bacteria (LAB) during fermentation and microbial composition in inasua. MATERIALS AND METHODS: The sample of inasua was taken from two traditional producers in Layeni village, Ceram Island. The diversity of lactic acid bacteria was analyzed based on the 16S rRNA gene sequence. RESULTS: The succession of lactic acid bacteria was strongly influenced by the physicochemical characteristics during fermentation. Lactobacillus plantarum was found dominant in both inasuas fermentation processes. At end of fermentation, L. plantarum was still found dominant in inasua with sap while inasua without sap was dominated by Leuconostoc mesenteroides. In addition, Lactobacillus paracasei (LAB) was found only in inasua with sap. The result of Denaturing Gradient Gel Electrophoresis (DGGE) revealed that Lactobacillus was the dominant bacteria in inasua with sap while Staphylococcus was dominant in inasua without sap. CONCLUSION: Inasua with sap was found with higher bacterial diversity index and lower evenness and dominance indices, as well as more complex LAB succession pattern during fermentation and bacterial composition, as opposed to inasua without sap.