Involvement of residues 296-299 in the enzymatic activity of tissue-type plasminogen activator

The tetra-alanine substitution variant KHRR 296–299 AAAAof tissue-type plasminogen activator (t-PA) was previously shownto have enhanced fibrin specificity and enhanced activity inthe presence of fibrin compared with the wild-type form of themolecule. The structural requirements for these alterationsin enzymatic activity were investigated by constructing severalamino acid substitution variants at each of the positions from296 to 299 and evaluating their activities under a variety ofconditions. Effects on plasminogen activator activity were commonamong the point mutants at positions 296–299; nearly allhad a phenotype similar to the KHRR 296–299 AAAA variant.The greatest effects on enzymatic function were found with multiplesubstitution variants, but some single charge reversals andproline substitutions had substantial effects. The enhancedfibrin specificity of KHRR 296–299 AAAA t-PA results inless fibrinogenolysis than seen with wild-type t-PA. Approximatelyfour times greater concentration of KHRR 296–299 AAAAcompared with wild-type t-PA was required to consume 50% ofthe fibrinogen in human plasma.     

Biomass estimation in batch biotechnological processes by Bayesian Gaussian process regression

This paper proposes a biomass concentration estimator for a batch biotechnological process based on Bayesian regression with Gaussian process. On the basis of experimental data, a two-stage bootstrap technique has been developed for the estimator design. In the first stage, the biomass data set was augmented with virtual filtered measurements, and in the second stage, the biomass estimator design was completed. The method provides information on the confidence level of the estimates, and the biomass estimator performances are illustrated for the Bacillus thuringiensis δ-endotoxins production process.     

Species,roasting degree and decaffeination influence the antibacterial activity of coffee against Streptococcus mutans

Coffee beverage has been associated with antibacterial activity against Streptococcus mutans, a cariogenic bacterium. This study aimed at identifying natural compounds in coffee that contribute to such activity and investigate the influence of species, roasting and decaffeination on it. Coffee chemical compounds and aqueous extracts of green and roasted regular and decaffeinated Coffea arabica and Coffea canephora beans were tested. MIC, biofilm inhibition and biofilm reduction results were correlated with the concentration of coffee compounds in the extracts. 5-Caffeoylquinic acid, trigonelline and caffeic acid solutions showed bacteriostatic activity (MIC = 0.8 mg/mL). Lighter and regular extracts showed higher inhibitory activity than darker and decaffeinated extracts, with an inverse correlation between bacterial colony-forming units and roasting degree. Only regular C. canephora extracts showed biofilm formation inhibition. The joint effect of chlorogenic acids, trigonelline and caffeine or other compounds removed by decaffeination seems to be one of the causes for coffee antibacterial activity against S. mutans.     

Chemical composition of a soybean cultivar lacking lipoxygenases (LOX2 and LOX3)

The nutrient, phytic acid, oxalate, trypsin inhibitors and isoflavones composition of a whole soy flour produced from a new cultivar (UFV-116), lacking lipoxygenases 2 and 3, compared to a conventional cultivar (OCEPAR-19) were determined. Protein and dietary fibres (total, soluble and insoluble) were similar for both cultivars. OCEPAR-19 was higher in lipids and UFV-116 in ash content (p < 0.05). Indispensable, dispensable and total aminoacid as well as Ca, K and Mg were higher for UFV-116. This cultivar also showed higher levels of phytic acid, oxalate and trypsin inhibitors (p < 0.05). Total saturated and unsaturated fatty acids were similar between them. However, palmitic and linoleic acids were higher for UFV-116 and stearic, α-linolenic and oleic acids for OCEPAR-19 (p < 0.05). The higher concentration of isoflavones in UFV-116 (p < 0.05) could provide better benefit for human health. Experimental studies are necessary to evaluate health effects of this new soybean cultivar.     

Evaluation of Norrish’s Equation for Correlating the Water Activity of Highly Concentrated Solutions of Sugars,Polyols, and Polyethylene Glycols

Norrish’s equation, a_\textw = X_\textw exp( - KX²_\texts )a_{{\text{w}}} = X_{{\text{w}}} \exp {\left( { - KX^{2}_{{\text{s}}} } \right)}, where a _w is water activity, X _w and X _s are molar fractions of water and solute, respectively, and K is the correlating constant, has been widely used to predict a _w of aqueous nonelectrolyte solutions in connection with development of intermediate moisture foods, i.e., food having a _w ≥ 0.85. Present work evaluated the ability of Norrish’s equation to model the water activity of solutions of sugars, polyols, and some polyethylene glycols, in a wide range of concentration, i.e., from low to highly concentrated solutions. For sugar and polyols, a relatively small modification of the “most accepted” literature parameters K allowed the fitting of the data for the wide range of solute concentrations corresponding to a range of a _w from 0.99 to about 0.3 for same solutes. However, a modified Norrish’s model needs to be used to model the behavior of polyethylene glycols 400 and 600 up to water activities as low as 0.5.     

Aflatoxins and ochratoxin A in Brazilian paprika

Seventy paprika samples collected in the city of São Paulo, Brazil, from January to April 2006 were analysed for aflatoxins and ochratoxin A (OTA) using an immunoaffinity column clean-up and HPLC-FLD. For aflatoxins, the limit of quantification (LOQ) were 0.23, 0.23, 0.45 and 0.45 μg/kg for AFB₁, AFB₂, AFG₁ and AFG₂, respectively. For OTA the LOQ was 0.80 μg/kg. Aflatoxins were found in 82.9% of samples and AFB₁ was detected in 61.4% at levels ranging from 0.5 to 7.3 μg/kg with mean concentration of 3.4 μg/kg. OTA was found in 85.7% at levels ranging from 0.24 to 97.2 μg/kg with mean concentration of 7.0 μg/kg.     

G-Quadruplex in Gene Encoding Large Subunit of Plant RNA Polymerase II: A Billion-Year-Old Story

G-quadruplexes have long been perceived as rare and physiologically unimportant nucleic acid structures. However, several studies have revealed their importance in molecular processes, suggesting their possible role in replication and gene expression regulation. Pathways involving G-quadruplexes are intensively studied, especially in the context of human diseases, while their involvement in gene expression regulation in plants remains largely unexplored. Here, we conducted a bioinformatic study and performed a complex circular dichroism measurement to identify a stable G-quadruplex in the gene RPB1, coding for the RNA polymerase II large subunit. We found that this G-quadruplex-forming locus is highly evolutionarily conserved amongst plants sensu lato (Archaeplastida) that share a common ancestor more than one billion years old. Finally, we discussed a new hypothesis regarding G-quadruplexes interacting with UV light in plants to potentially form an additional layer of the regulatory network.     

A Beckwith–Wiedemann-Associated CDKN1C Mutation Allows the Identification of a Novel Nuclear Localization Signal in Human p57Kip2

p57^Kip2 protein is a member of the CIP/Kip family, mainly localized in the nucleus where it exerts its Cyclin/CDKs inhibitory function. In addition, the protein plays key roles in embryogenesis, differentiation, and carcinogenesis depending on its cellular localization and interactors. Mutations of CDKN1C, the gene encoding human p57^Kip2, result in the development of different genetic diseases, including Beckwith–Wiedemann, IMAGe and Silver–Russell syndromes. We investigated a specific Beckwith–Wiedemann associated CDKN1C change (c.946 C>T) that results in the substitution of the C-terminal amino acid (arginine 316) with a tryptophan (R316W-p57^Kip2). We found a clear redistribution of R316W-p57^Kip2, in that while the wild-type p57^Kip2 mostly occurs in the nucleus, the mutant form is also distributed in the cytoplasm. Transfection of two expression constructs encoding the p57^Kip2 N- and C-terminal domain, respectively, allows the mapping of the nuclear localization signal(s) (NLSs) between residues 220–316. Moreover, by removing the basic RKRLR sequence at the protein C-terminus (from 312 to 316 residue), p57^Kip2 was confined in the cytosol, implying that this sequence is absolutely required for nuclear entry. In conclusion, we identified an unreported p57^Kip2 NLS and suggest that its absence or mutation might be of relevance in CDKN1C-associated human diseases determining significant changes of p57^Kip2 localization/regulatory roles.     

Chemical characterisation of non-defective and defective green arabica and robusta coffees by electrospray ionization-mass spectrometry (ESI-MS)

The coffee roasted in Brazil is considered to be of low quality, due to the presence of defective coffee beans that depreciate the beverage quality. These beans, although being separated from the non-defective ones prior to roasting, are still commercialized in the coffee trading market. Thus, it was the aim of this work to verify the feasibility of employing ESI-MS to identify chemical characteristics that will allow the discrimination of Arabica and Robusta species and also of defective and non-defective coffees. Aqueous extracts of green (raw) defective and non-defective coffee beans were analyzed by direct infusion electrospray ionization mass spectrometry (ESI-MS) and this technique provided characteristic fingerprinting mass spectra that not only allowed for discrimination of species but also between defective and non-defective coffee beans. ESI-MS profiles in the positive mode (ESI(+)-MS) provided separation between defective and non-defective coffees within a given species, whereas ESI-MS profiles in the negative mode (ESI(−)-MS) provided separation between Arabica and Robusta coffees.