TA-Cloning for Diabetes Treatment: Expressing Corynebacterium Malic Enzyme Gene in E. coli.

Diabetes mellitus represents a persistent metabolic condition marked by heightened levels of blood glucose, presenting a considerable worldwide health concern, and finding targeted treatment for it is a crucial priority for global health. Gram-positive aerobic bacteria, predominantly inhabiting water and soil, are known carriers of various enzyme-encoding genetic material, which includes the malic enzyme gene that plays a role in insulin secretion. Corynebacterium glutamicum bacteria (ATCC 21799) were acquired from the Pasteur Institute and confirmed using microbiological and molecular tests, including DNA extraction. After identification, gene purification and cloning of the maeB gene were performed using the TA Cloning method. Additionally, the enhancement of enzyme expression was assessed using the expression vector pET-28a, and validation of simulation results was monitored through a real-time PCR analysis. Based on previous studies, the malic enzyme plays a pivotal role in maintaining glucose homeostasis, and increased expression of this enzyme has been associated with enhanced insulin sensitivity. However, the production of malic enzyme has encountered numerous challenges and difficulties. This study successfully isolated the malic enzyme genes via Corynebacterium glutamicum and introduced them into Escherichia coli for high-yield production. According to the results, the optimum temperature for the activity of enzymes has been identified as 39 °C.

Antibacterial Property of Silver Nanoparticles Green Synthesized from Stachys schtschegleevii Plant Extract on Urinary Tract Infection Bacteria.

Urinary tract infections are one of the most common infections worldwide. Given the increasing antibiotic resistance, monitoring antibiotic sensitivity patterns is crucial. Furthermore, silver nanoparticles synthesized from Stachys schtschegleevii can exhibit potent antibacterial, antibiotic, and antifungal properties. The plant S. schtschegleevii was collected from its natural habitat, dried, and its extract was then exposed to silver nitrate. Under specific conditions, silver nanoparticles were synthesized from it. Subsequently, the production and validation of silver nanoparticles were confirmed through techniques such as FTIR analysis, UV-Vis analysis, TEM, SEM, EDX analysis, and zeta potential analysis. In the in vitro section of the research, the impact of the extracted silver nanoparticles on bacteria isolated from patients' urine and standard bacterial culture (control) was assessed using the disc diffusion and MIC test methods. The results of the analyses are FTIR (high protein content; proteins and phenols serve as stabilizing agents), UV-Vis (peak of 460 nm), TEM (spherical to occasionally elliptical shapes), SEM (sizes: 26 to 72 nm), EDX (peak at 3 keV), and zeta potential (- 15.76 ± 0.05 mV). The effect of silver nanoparticles by disc diffusion method (mm) is Enterococcus faecalis = 18.31 ± 0.35, Escherichia coli = 21.51 ± 0.61, and Staphylococcus aureus = 19.02 ± 1.28, and by MIC test (µg/ml), E. faecalis = 19, E. coli = 18, and Staphylococcus aureus = 16. Antibacterial activity of the silver nanoparticles synthesized from S. schtschegleevii means that these herbal nanoparticles treat urinary tract infections caused by some of the test isolates.