Immobilization of Bacillus Subtilis E6-5 Protease and Commercial Protease in Nanofibrils Containing Different Amino Acids

DergiPark: 633788trkjnatBu çalışmada, elektrospin yöntemiyle yüksek yüzey alanına sahip, glisin, tirozin ve glutamik asit aminoasitleri ile oluşturulmuş poliamid 6 polimer yüzeyler üretilmiş ve liyofilize Bacillus subtilis E6-5 proteaz ve ticari proteaz enzimleri nanofibriller üzerinde immobilize edilmiştir. Enzimlerin yeniden kullanılabilirliği araştırıldı. Enzimlerin immobilizasyon verimlilikleri yaklaşık olarak % 50-55 arasındaydı. Liyofilize Bacillus proteazı ile yapılan çalışmalarda glutaraldehitle aktifleştirilmiş PA6 nanolifler ve glutaraldehitle aktifleştirilmeyen PA6 nanoliflerde glutamik asit aminoasidi varlığında immobilizasyonun daha başarılı olduğu saptanmıştır. Glutaraldehit ile aktifleştirilmemiş ve aktifleştirilmiş yüzeylerde immobilize edilen liyofilize proteaz enziminin 4 kez kullanımı olmasına rağmen, en iyi işlevsel stabilite 2 kez kullanım ile elde edilmiştir. Saf PA6/glutamik asit nanoliflerinde iki tekrarlı kullanım sonucu enzimin immobilizasyon verimi % 38 olarak bulunmuştur. Glutaraldehitle aktifleştirilmiş PA6 nanoliflerde de PA6/glutamik asit nanolif yüzeyleri iki tekrarlı kullanım sonucu enzimin immobilizasyon verimi % 65 olarak bulunmuştur. Nanoliflerin glutaraldehitle aktifleştirmesi sonucu enzim immobilizasyon verimi iki kat arttırılmıştır. Ticari proteaz ile yapılan çalışmalarda ise glutaraldehitle aktifleştirilmemiş nanolif yüzeylerde enzimin 6 kez kullanımı olmasına rağmen en işlevsel stabilite 3 tekrarlı kullanımda elde edilmiştir. En başarılı immobilizasyon verimi PA6 nanoliflerde % 58 olarak bulunmuştur. Glutaraldehitle aktifleştirilmiş PA6 nanoliflerde de enzim 6 kez kullanım bulmuş fakat işlevsel stabilite 4 tekrarlı kullanıma kadar korunmuştur.In this study, polyamide 6 polymer surfaces that have a high surface area were produced by electrospinning method with the participation of Glycine, Tyrosine and Glutamic acid amino acids, and lyophilized Bacillus subtilis E6-5 protease and commercial protease enzymes were immobilized on nanofibrils. Enzyme reusability were investigated. The immobilization efficiencies of the enzymes were approximately between 50-55 %. In studies with lyophilized Bacillus protease, glutaraldehyde activated PA6 nanofibrils and glutaraldehyde unactivated PA6 nanofibrils were found to be more immobilized in the presence of Glutamic acid. Although the lyophilized protease enzyme immobilized on non-glutaraldehyde activated and activated surfaces has been used 4 times, the best functional stability has been achieved with 2 times use. In pure PA6/Glutamic acid nanofibrils, the immobilization yield of the two times used enzymes was found to be 38 %. In glutaraldehyde-activated PA6 nanofibrils, the PA6/Glutamic acid nanofibril surfaces were found to have 65 % immobilization yield of the two repetitive used enzymes. The enzyme immobilization efficiency has been doubled by glutaraldehyde activation of the nanofibrils. In studies with commercial protease, the most functional stability was obtained for 3 repeated uses, although the enzyme was used 6 times on the non-glutaraldehyde activated nanofibril surfaces. The most successful immobilization was found in 58 % of PA6 nanofibrils. In glutaraldehyde-activated PA6 nanofibrils, the enzyme was found to be used 6 times, but the functional stability was maintained as much as 4 times of repeated use

Inhibition of Pseudomonas aeruginosa biofilm formation and motilities by human serum paraoxonase (hPON1)

Human serum paraoxonase 1 (hPON1) which hydrolyzes Pseudomonas aeruginosa acyl homoserine lactone (AHL) signal molecules was used as antibiofilm agent. hPON1 was purified by using ammonium sulfate precipitation and specially designed hydrophobic interaction chromatography (Sepharose 4B-L-tyrosine-1-Naphthylamine) from the fresh human serum. As cell motility of swarming, swimming and twitching are proven instrumental in biofilm formation, we investigated whether or not hPON1 affected the P. aeruginosa motility. hPON1 was reduced the early stage of biofilm formation, mature biofilm and motilities. The early stage and old biofilm were decreased more than 50% by 1 mg ml–1 of hPON1 concentration within range of 0.1–10 mg ml–1. Additionally, exopolymeric substance (EPS) of mature biofilm was indirectly decreased by hPON1. Inhibitory effect of hPON1 within range of 0.003–30 mg ml–1 on swarming and swimming motilities. But it resulted in highly inhibitory effects on twitching motility at concentration as low as 0.3 mg ml–1 concentration. This study proved that hPON1 alone can be safely used to inhibit/disrupt the mature biofilms and cell motility of P. aeruginosa and beholds much promise in clinical applications

Serious gaming in cbrne domain: a survey on user expectations, concerns and suggestions

Serious gaming, the umbrella term describing the video and board games having an ulterior motive rather than only entertainment, is widely used in several domains such as health, education, and military. The use of serious gaming in Chemical, Biological, Radiological, Nuclear, and Explosives (CBRNe) domain is quite a new concept, which makes it open to several questions, misunderstandings (e.g. game or simulation), and expectations. In this study, a pre-development survey for a serious game on CBRNe [1], which is currently been developed for European Network Of CBRN TraIning CEnters (eNOTICE) project [2], is presented. The serious game in eNOTICE aims to train the CBRNe personnel under different scenarios, conditions, and environments. Before the game is developed and adapted to different scenarios, we formed a 24-question questionnaire [3] under three sections for the purposes of: 1) collecting information on participant’s video-gaming background, 2) evaluating participant’s idea on Serious Games / Games for Change, and 3) assessing user expectations from eNotice Serious Game. For the time being, 14 CBRNe professionals from different backgrounds participated to the survey, answered the questions, and filled the open-ended forms on suggestions. The analysis of the results demonstrates a preliminary layout of opportunities and challenges laying ahead in this domain with a list of the main expectations, misconceptions and suggestions. Majority of the participants (13 out of 14 participants) was highly positive about pursuing CBRNe training using serious games while the only person giving negative feedback was against using video games in general, but still suggested how to integrate them to the training—which is a positive step towards idea exchange and brainstorming. This preliminary study demonstrates the fact that the use of serious gaming in CBRNe domain is a promising and expected research concept which will further enhance the capabilities of training centers. Acknowledgments: This work is supported by European Network Of CBRN TraIning Centers (eNOTICE) project funded under EU H2020 (Project ID: 740521)

Optimization of culture medium for the production and partial purification and characterization of an antibacterial activity from brevibacillus laterosporus strain EA62

The present study was performed to report the bacterial identification, optimization of culture conditions and characterization of a novel antibiotic substance from a Bacillus sp. EA62 strain isolated from soil. The EA62 strain was identified based on 16S rRNA analysis. The new isolate EBD 9-1 showed 100% sequence identity with Brevibacillus laterosporus. The antibacterial activity of the new substance was examined against five pathogenic bacteria and was observed to be the most effective against Escherichia coli. An agar diffusion assay was performed to evaluate the antibacterial activity of the substance. The effects of some nutritional (amino acid, carbon, nitrogen and metal sources) and physical factors (pH and temperature) and incubation time on the antibiotic activity were studied. Antibiotic activity in basal medium reached the maximum levels after 72 h of incubation. The best antibiotic activity was obtained in the presence of glucose as a carbon source, yeast extract as a nitrogen source, glutamic acid as an amino acid source and MgSO4+CaCO3 as metal ion sources. For physical parameters, the best results were obtained at 37 degrees C, pH 7.0. The antibiotic substance was partially purified, and the estimated molecular weight was 6.3 kDa. The minimum inhibitory concentration (MIC) values determined against five pathogen bacteria were >256 mu g/ml. The substance was identified by thin-layer chromatography, and its Rf value was measured as 0.04 cm

Evaluation of Glutathione S-Transferase P1 Polymorphisms (Ile105Val and Ala114Val) in Patients with Small Cell Lung Cancer

Aims: Glutathione S-transferase P1 (GSTP1) plays an important role in cellular protection against oxidative stress and toxic chemicals. Polymorphisms within GSTP1 are associated with alterations in enzyme activity, which may lead to development of lung disease and cancer. In this study, we aimed to investigate the GSTP1 Ile105Val and Ala114Val polymorphisms in patients with small cell lung cancer (SCLC). Patients/Methods: GSTP1 Ile105Val polymorphism in exon 5 and GSTP1 Ala114Val polymorphism in exon 6 were determined by using polymerase chain reaction-restriction fragment length polymorphism techniques in 89 patients with SCLC and 108 control patients with chronic obstructive pulmonary disease (COPD). Genotype frequencies and cigarette smoking intensities were compared among SCLC and COPD patients. Results: There were significantly less SCLC patients with variant exon 6 genotypes than COPD patients (7.9% vs. 20.4%, p = 0.007), while the number of patients with variant exon 5 genotypes were comparable among groups. SCLC and COPD patients with variant exon 6 genotype showed trends toward exhibiting reduced cigarette consumption. Conclusions: The variant GSTP1 exon 6 genotype might be conferring protection against SCLC development. Whether this effect is associated with exposure to cigarette smoking needs to be clarified

Evaluation of Glutathione S-Transferase P1 Polymorphisms (Ile105Val and Ala114Val) in Patients with Small Cell Lung Cancer

Aims: Glutathione S-transferase P1 (GSTP1) plays an important role in cellular protection against oxidative stress and toxic chemicals. Polymorphisms within GSTP1 are associated with alterations in enzyme activity, which may lead to development of lung disease and cancer. In this study, we aimed to investigate the GSTP1 Ile105Val and Ala114Val polymorphisms in patients with small cell lung cancer (SCLC). Patients/Methods: GSTP1 Ile105Val polymorphism in exon 5 and GSTP1 Ala114Val polymorphism in exon 6 were determined by using polymerase chain reaction-restriction fragment length polymorphism techniques in 89 patients with SCLC and 108 control patients with chronic obstructive pulmonary disease (COPD). Genotype frequencies and cigarette smoking intensities were compared among SCLC and COPD patients. Results: There were significantly less SCLC patients with variant exon 6 genotypes than COPD patients (7.9% vs. 20.4%, p = 0.007), while the number of patients with variant exon 5 genotypes were comparable among groups. SCLC and COPD patients with variant exon 6 genotype showed trends toward exhibiting reduced cigarette consumption. Conclusions: The variant GSTP1 exon 6 genotype might be conferring protection against SCLC development. Whether this effect is associated with exposure to cigarette smoking needs to be clarified