Lalida Shank

Screening of filamentous fungi for production of myrosinase

A linhagem Aspergillus sp. NR46FB, isolada de solo atraves da tecnica do agar sulfato de bario-sinigrina, foi testada quanto a producao de mirosinase. O fungo degradou completamente o glicosinolato e produziu 3,19 U.mL-1 de mirosinase, apos 48 h de cultivo. Devido a alta producao de mirosinase, esse novo isolado e um potente candidato para aplicacoes industriais.

GM2-Activator Protein: A New Biomarker for Lung Cancer

Introduction: Effective biomarkers for early diagnosis of lung cancer are needed. A recent study demonstrated that urinary GM2-activator protein (GM2AP) level was increased in lung cancer patients. This study aims to validate the potential application of GM2AP as a biomarker for diagnosis of lung cancer. Methods: Serum and urine samples were obtained from 189 participants (133 patients for treatment naive lung cancer, 26 healthy volunteers for urine, and 30 healthy volunteers for serum). GM2AP level was detected by Western blotting and quantified using enzyme-linked immunosorbent assay (ELISA). The GM2AP expression in tumors and nontumor parts of lung tissues from 143 nonsmall cell lung cancers was detected by immunohistochemical stains. Results: There was an 8.11 ± 1.36 folds increase in urine and a 5.41 ± 0.73 folds increase in serum level of GM2AP in lung cancer patients compared with healthy volunteers (p < 0.0001), achieving a 0.89 AUC value in urine and 0.90 AUC value in serum for the receiver-operating characteristic curves. Both serum and urine levels of GM2AP correlated significantly with pathology stages (urine, p = 0.009; serum, p < 0.0001). Using immunohistochemical, positive expression of GM2AP was found at 83.9% of nonsmall cell lung cancers patients and none in normal tissue. The GM2AP expression was significantly correlated with pathology stage (p = 0.0001). Patients with higher GM2AP expression had shorter overall survival (p = 0.045) and disease-free survival (p = 0.049) than lower GM2AP expression. Moreover, the multivariate analysis suggested GM2AP as an independent predictors of disease-free survival and overall survival. Conclusions: Our study demonstrates that GM2AP might serve as potential diagnostic and prognostic biomarkers in patients with lung cancer.

Functional importance of the Gly cluster in transmembrane helix 2 of the Bordetella pertussis CyaA-hemolysin: Implications for toxin oligomerization and pore formation

Adenylate cyclase-hemolysin (CyaA) is a major virulence factor of Bordetella pertussis causing whooping cough in humans. We previously showed that two transmembrane helices (α2 and α3) in the hemolysin domain (CyaA-Hly) are crucially involved in hemolytic activity. Here, PCR-based substitutions were employed to investigate a potential involvement in hemolysis of a series of four Gly residues (Gly(530), Gly(533), Gly(537) and Gly(544)) which map onto one face of a helical wheel plot of pore-lining helix 2. All CyaA-Hly mutant toxins were over-expressed in Escherichia coli as 126-kDa soluble proteins at levels comparable to the wild-type toxin. A drastic reduction in hemolytic activity against sheep erythrocytes was observed for three CyaA-Hly mutants, i.e. G530A, G533A and G537A, but not G544A, suggesting a functional importance of the Gly(530)_Gly(533)_Gly(537) cluster. A homology-based structure of the α2-loop-α3 hairpin revealed that this crucial Gly cluster arranged as a GXXGXXXG motif is conceivably involved in helix-helix association. Furthermore, a plausible pore model comprising three α2-loop-α3 hairpins implicated that Gly(530)XXGly(533)XXXGly(537) could function as an important framework for toxin oligomerization. Altogether, our present data signify for the first time that the Gly(530)_Gly(533)_Gly(537) cluster in transmembrane helix 2 serves as a crucial constituent of the CyaA-Hly trimeric pore structure.

Fabrication of Ascorbic Acid Biosensor Based on Coupling Polyethylene Glycol Modified Silk Fibroin Membrane onto Glassy Carbon Electrode

Nowadays biosensors have been extensively used in a wide variety of applications especially in clinical works and food industry. In this work, a specific ascorbic acid (AA) biosensor was developed by immobilizing ascorbate oxidase (ASOD) on a polyethylene glycol (PEG) modified silk fibroin (SF) membrane then coupling to the glassy carbon electrode (GCE). The SF-PEG-ASOD membrane provided the highest enzyme activity in phosphate buffer at pH 5. As being the electrode, the SF-PEG-ASOD modified GCE displayed the highest response when it is operated under the condition of 0.40 mg/L of ASOD in phosphate buffer at pH 5. This biosensor provided both good linearity (r2 = 0.999 in the range of 1.0-10.0 mM) and sensitivity with short response time (26s). It also exhibited good anti-interference ability with the storage time of 5 days without changing its initial response.