Biomolecular engineering最新文献

The present investigation makes a comparative investigation of individual light source on the different commercially important pigments in Spirulina fussiformis in photobioreactor culture condition. Continuous culture system was carried out throughout the experimental condition. Initially, seed culture, corresponding to 0.2 g/L on dry weight basis was cultivated in Zarrouks medium with different colored light source in reactor. Maximum daily biomass productivity, 0.8 g/L, 0.75 g/L and 0.69 g/L in white light (WL), blue light (BL) and green light (GL), respectively, conditions was noticed. Pigment content during WL treatment showed the highest accumulation (5.5 μg/mL) of chlorophyll whereas, other pigments roughly remained constant without much change, implying WL intensity is better for chlorophyll synthesis. On the other hand, chlorophyll and phycocyanin content gradually increased up to 7 μg/mL and 2 mg/mL, respectively, at BL intensity. The response to GL was negative to all pigments studied except for phycocyanin; in this case a highest production (2.5 mg/mL) was seen during 18 days experimental period. Additionally, when yellow light (YL) treatment experiments were conducted, the rate of production gradually decreased from 6th day onward in all pigments demonstrating the photobleaching effect of YL. The average rate of pigments production did not show significant accumulation in red light (RL) light treatment except phycoerythrin which showed an increasing trend of production.

It is worth to mention here that higher light intensity is better for production of phycocyanin and phycoerythrin in Spirulina.

RNA sequences can form structures which are conserved throughout evolution and the question of aligning two RNA secondary structures has been extensively studied. Most of the previous alignment algorithms require the input of gap opening and gap extension penalty parameters. The choice of appropriate parameter values is controversial as there is little biological information to guide their assignment. In this paper, we present an algorithm which circumvents this problem. Instead of finding an optimal alignment with predefined gap opening penalty, the algorithm finds the optimal alignment with exact number of aligned blocks.

Myocilin (MYOC, TIGR) variations are associated with juvenile and primary open angle glaucoma (POAG). To investigate consequences of MYOC wildtype overexpression and selected mutations, we established a heterologous insect cell system (High Five™). Wildtype, Pro370Leu, Gln368X and Lys423Glu were cloned into a modified pIB/V5-His (pEXIV) vector with and without downstream GFP in frame fusion. Mutations were introduced by in vitro mutagenesis. Heterologous expression was shown and analysed by RT-PCR, Western blotting, immunocytochemistry and fluorescence microscopy. Extended cultivation (>14 days) resulted in accumulation of MYOC protein for all variants in growing dilated cisterns of the rough endoplasmic reticulum. Finally cell death for overexpressed wildtype and mutants occurs. A direct attachment of ribosomes to these growing vesicles preceding the cell death was observed by electron microscopy. Our observations indicate that this system is suitable to trace the intracellular effects of MYOC mutants.

Metabolic pathway databases such as KEGG contain information on thousands of biochemical reactions drawn from the biomedical literature. Ensuring consistency of such large metabolic pathways is essential to their proper use. In this paper, we present a new method to determine consistency of an important class of biochemical reactions. Our method exploits the knowledge of the atomic rearrangement pattern in biochemical reactions, to reduce the automatic atom mapping problem to a series of chemical substructure searches between the substrate and the product of a biochemical reaction. As an illustrative application, we describe the exhaustive validation of a substantial portion from the latest release of the KEGG LIGAND database.

In-stent restenosis is a process that occurs in 10–50% of cases currently treated with stent and it is caused by an abnormal smooth muscle cell (SMC) proliferation and migration in the vascular lumen. One of the most promising strategy to reduce restenosis is stent coating with biodegradable polymers to deliver in situ anti-proliferative drugs. Poly(d,l)lactic acid (P(d,l)LA), one of the most interesting candidate for stent coating, has been observed to induce inflammation and neointimal proliferation. In our laboratory, we developed P(d,l)LA enriched with Vitamin E (Vit.E), an anti-oxidative and anti-inflammatory agent that reduces also SMC proliferation. In order to evaluate the in vitro cellular behaviour of neointima cells onto Vitamin E-enriched P(d,l)LA, cell adhesion and proliferation along with the expression of two SMC migration markers (MMP-9 and hyaluronic acid receptor CD44) were measured in rat vascular SMC A10 cells seeded onto control P(d,l)LA (PLA) and P(d,l)LA films containing 10–30% (w/w) Vit.E (PLA10–30). Cell adhesion, proliferation and MMP-9 production were unaffected by the Vit.E presence in the PLA films after 24 h, while proliferation was slowed or blocked after 48 and 72 h onto PLA10, 20 and 30. MMP-9 production was almost blocked and CD44 density decreased significantly after 72 h for cells grew onto PLA30 compare to cells seeded onto PLA. These data indicate that Vit.E-enriched P(d,l)LA could be an interesting polymer for stent coating.

Due to its strength and specificity, the interaction between avidin and biotin has been used in a variety of scientific and medical applications ranging from immunohistochemistry to drug targeting. The present study describes two methods for biotinylation of proteins secreted from eukaryotic cells using the Escherichia coli biotin protein ligase. In one system the biotin ligase was co-secreted from the cells along with substrate protein enabling extracellular biotinylation of the tagged protein. In the other system, biotin ligase was engineered to be retained in the endoplasmic reticulum (ER) and metabolically biotinylates the secretory protein as it passes through the ER. An engineered antibody fragment, a diabody with specificity for carcinoembryonic antigen (CEA) was fused to the biotin acceptor domain (123 amino acid) of Propionibacterium shermanii. Coexpression of the fusion protein with ER retained biotin ligase showed higher biotinylation efficiency than biotinylation by co-secreted ligase. Biotinylation of the anti-CEA diabody tagged with a short (15 amino acid, Biotin Avitag™) biotin acceptor peptide was also successful. Utilization of ER retained biotin ligase for biotinylation of protein is an attractive alternative for efficiently producing uniformly biotinylated recombinant proteins for a variety of avidin–biotin technologies.

A peak is a pair of real values $\left(x\text{,}y\right)$, where x is the time when peak of height y is registered. In the peak alignment problem, we are given two sequences of peaks, and our task is to align the sequences allowing some basic edit operations on the peaks. We study an instance of the peak alignment problem that arises in the analysis of Mass Spectrometry data in Systems Biology. There the measurement technique guarantees that two peaks $\left(x\text{,}y\right)$, $\left(x^{\prime }\text{,}{y}^{\prime }\right)$ can only be considered the same if x is close enough to $x^{\prime }$, and y is close enough to $y^{\prime }$. We review some methods to do alignment under such restrictions on matches.

Recent developments in the study of RNA silencing indicate that double-stranded RNA (dsRNA) can be used in eukaryotes to block expression of a corresponding cellular gene. There is also a large class of small non-coding RNAs having potential to form a distinct, stable stem-loop in numbers of eukaryotic genomes. We had reported that a large imperfect dsRNA structure with hundreds of base-pairs (bp) in the $3^{\prime }$ untranslated region ($3^{\prime }$UTR) of cytotoxic ribonuclease was correlated with the translation suppression. In this study, we search for such dsRNAs in a $3^{\prime }$ UTR database. The occurrence rate of large dsRNA in $3^{\prime }$ UTRs ranges from 0.01% in plant to 0.30% in vertebrate mRNAs. However, small imperfect dsRNAs of $\sim$ 30 bp are much more prevalent than large ones. The small dsRNAs are statistically very significant and uniquely well-ordered. Most of them have the conserved structural features of pre-miRNAs. Our data mining of the dsRNAs in the $3^{\prime }$ UTR database can be used to explore RNA-based regulation of gene expression.

Aptamers are single stranded DNA or RNA ligands which can be selected for different targets starting from a huge library of molecules containing randomly created sequences. Aptamers have been selected to bind very different targets, from proteins to small organic dyes.

In addition to the very important aspect of having an unlimited source of identical affinity recognition molecules available due to the selection process, aptamers can offer advantages over antibodies that make them very promising for analytical applications. The use of aptamers as therapeutic tools is nowadays well established. On the contrary, the analytical application of aptamers in diagnostic devices or in systems for environmental and food analysis, is still under investigation and the scientific community still need further research to demonstrate the advancements brought by this new kind of ligands.

This review will focus on these latter applications with particular attention to the detection of food pathogens, terrorism threat agents, thrombin and cytokines.

A human interferon beta (hINF-β) synthetic gene was optimized and expressed in Escherichia coli BL21-SI using a vector with the T7 promoter. To determine the best culture conditions such as culture medium, temperature, cell density and inducer concentration, we used the response surface methodology and a Box-Behnken design to get the highest hINF-β production. The maximum hINF-β production of 61 mg l⁻¹ was attained using minimum medium and the following predicted optimal conditions: temperature of 32.5 °C, cell density of 0.64, and inducer concentration of 0.30 M NaCl. This is the first report showing the successful performance of the BL21-SI system in a minimum medium. The response surface methodology is effective for the optimization of recombinant protein production using synthetic genes.