Dorota Kowalska

Synthetic Small-Molecule Prohormone Convertase 2 Inhibitors

The proprotein convertases are believed to be responsible for the proteolytic maturation of a large number of peptide hormone precursors. Although potent furin inhibitors have been identified, thus far, no small-molecule prohormone convertase 1/3 or prohormone convertase 2 (PC2) inhibitors have been described. After screening 38 small-molecule positional scanning libraries against recombinant mouse PC2, two promising chemical scaffolds were identified: bicyclic guanidines, and pyrrolidine bis-piperazines. A set of individual compounds was designed from each library and tested against PC2. Pyrrolidine bis-piperazines were irreversible, time-dependent inhibitors of PC2, exhibiting noncompetitive inhibition kinetics; the most potent inhibitor exhibited a Ki value for PC2 of 0.54 μM. In contrast, the most potent bicyclic guanidine inhibitor exhibited a Ki value of 3.3 μM. Cross-reactivity with other convertases was limited: pyrrolidine bis-piperazines exhibited Ki values greater than 25 μM for PC1/3 or furin, whereas the Ki values of bicyclic guanidines for these other convertases were more than 15 μM. We conclude that pyrrolidine bis-piperazines and bicyclic guanidines represent promising initial leads for the optimization of therapeutically active PC2 inhibitors. PC2-specific inhibitors may be useful in the pharmacological blockade of PC2-dependent cleavage events, such as glucagon production in the pancreas and ectopic peptide production in small-cell carcinoma, and to study PC2-dependent proteolytic events, such as opioid peptide production.

Modulation of PC1/3 activity by self-interaction and substrate binding.

Prohormone convertase (PC)1/3 is a eukaryotic serine protease in the subtilase family that participates in the proteolytic maturation of prohormone and neuropeptide precursors such as proinsulin and proopiomelanocortin. Despite the important role of this enzyme in peptide synthesis, how PC1/3 activity is regulated is still poorly understood. Using ion exchange chromatography and two-dimensional gel electrophoresis we found that natural PC1/3 present in AtT-20 cells and bovine chromaffin granules, as well as recombinant PC1/3 secreted from overexpressing Chinese hamster ovary cells, exists as multiple ionic forms. Gel filtration and cross-linking studies revealed that protein oligomerization and aggregation contribute greatly to variability in surface charge. The most acidic forms of PC1/3 contained both inactive aggregates as well as oligomerized 87-kDa PC1/3 that exhibited stable activity which was partially latent and could be revealed by dilution. No such latency was observed for the more basic, 66/74-kDa forms of PC1/3. Fractions containing these species were stabilized by preincubation with micromolar concentrations of either fluorogenic substrate or peptides containing pairs of basic residues. In addition, the most active form of 87-kDa PC1/3, a probable homodimer, was activated by preincubation with these same peptides. Cleavage by PC1/3 is often the initiating step in the biosynthetic pathway for peptide hormones, implying that this is a natural step for regulation. Our data suggest that enzyme oligomerization and peptide stabilization represent important contributing factors for the control of PC1/3 activity within secretory granules.

Characterization of the essential oil from cone-berries of Juniperus communis L. (Cupressaceae)

Summary Juniperus communis L. (Cupressaceae) is a plant widely cultivated in the Northern hemisphere. Juniper berries, the fruit of Juniperus communis L. are a highly valued, essential oil-rich plant material used traditionally in folk medicine as antiseptic, diuretic, antirheumatic, anti-inflammatory, antibacterial and antifungicidal agent. This paper reviews information on extraction methods of the essential oil from the juniper berries, its chemical composition and antimicrobial as well as antioxidant properties.

Acrylamide in food products – eating habits and consumer awareness among Medical School students

INTRODUCTION Acrylamide is formed in several foods during high-temperature processing. In view of reports written about the neurotoxic, genotoxic and carcinogenic effects of acrylamide, it was considered that the presence of this substance in food products might pose a risk for human health. Currently, according to EU Commission recommendations, the content of acrylamide in food should be monitored. OBJECTIVE The aim of this work was to analyze the food preferences of youth and students from medical schools in Radom, central-eastern Poland, as the most frequent precipitantsas in the field of food products that may be a significant source of acrylamide in the diet. Furthermore, an attempt was made to determine the level of knowledge of the population in the field of acrylamide. MATERIAL AND METHODS The research was conducted by questionnaire. The study was based on the answers of 227 respondents. The survey was carried out by direct contact with an interviewer from February - June 2012. RESULTS Analysis of the study population shows that women consume more coffee than men. In addition, adults over 25 years old consumed the largest quantity of coffee; it can therefore be assumed that it is a significant source of acrylamide in their bodies. However, even young people under 17 declared that they consume coffee every day (20%). CONCLUSIONS Due to the adverse effects of this compound it is important to reduce the level of acrylamide in food products. A few people in the population (7%) had heard of acrylamide previously, but none of them had any knowledge of its occurrence and formation. It is necessary to take strong action to change attitudes towards acrylamide and attempt to introduce ways to reduce this compound in the diet, for example, by appropriate selection of products in the daily diet and appropriate means of thermal preparation of products at home.