Ionic Liquid-Based Periodic Mesoporous Organosilica: An Innovative Matrix for Enzyme Immobilization

Employment of enzyme immobilization technique for α-amylase, provides useful approaches to improve enzyme resistance to harsh conditions and reusability. Two silica supports, Ionic Liquid-Based Periodic Mesoporous Organosilica which we call it PMO-IL, and SBA-15, have been used for immobilization of α-amylase. We found that proper electrostatic interactions result in a higher immobilization yield of PMO-IL rather than SBA-15. The kinetics parameters show that α-amylase@PMO-IL activity has slightly decreased in comparison to free enzyme and the stability studies indicate that it is even more active than free enzyme in the natural pH which shows the improving role of ionic liquid on enzyme structure and function. Also, both immobilized enzymes have 2 times more stability at 70 °C and 80 °C after 60 min in comparison to free enzyme. Finally, the recovery efficiency of α-amylase@PMO-IL is 88% of its initial activity after 4 cycles which is 23% more than a previous report

Ionic Liquid-Based Periodic Mesoporous Organosilica: An Innovative Matrix for Enzyme Immobilization

Employment of enzyme immobilization technique for α-amylase, provides useful approaches to improve enzyme resistance to harsh conditions and reusability. Two silica supports, Ionic Liquid-Based Periodic Mesoporous Organosilica which we call it PMO-IL, and SBA-15, have been used for immobilization of α-amylase. We found that proper electrostatic interactions result in a higher immobilization yield of PMO-IL rather than SBA-15. The kinetics parameters show that α-amylase@PMO-IL activity has slightly decreased in comparison to free enzyme and the stability studies indicate that it is even more active than free enzyme in the natural pH which shows the improving role of ionic liquid on enzyme structure and function. Also, both immobilized enzymes have 2 times more stability at 70 °C and 80 °C after 60 min in comparison to free enzyme. Finally, the recovery efficiency of α-amylase@PMO-IL is 88% of its initial activity after 4 cycles which is 23% more than a previous report

Journal of Petroleum Science and Technology *Corresponding author THE GENE CLONING, OVEREXPRESSION, PURIFICATION, AND CHARACTERIZATION OF DIBENZOTHIOPHENE MONOOXYGENASE AND DESULFINASE FROM GORDONIA ALKANIVORANS RIPI90A

ABSTRACT The biodesulfurization (BDS) of sulfur compounds in fossil fuels is a process to reduce sulfur dioxide emissions that cause environmental pollution. Gordonia alkanivorans RIPI90A is able to convert dibenzothiophene, an organic sulfur compound in petroleum, to 2-hydroxybiphenyl (2-HBP) in 4S pathway. In this study, (DBT), DszA and DszB, DBT sulfone monooxygenase, and desulfinase were respectively isolated from G. alkanivorans RIPI90A. PCR amplified fragments were obtained by using primers designed based on known sequences from G. alkanivorans RIPI90A. They are identified as dszA and dszB and have shown high similarity compared to Rhodococcus erythropolis IGTS8 (88% for dszA and 88% for dszB). Subsequently, dszA and dszB genes were expressed under the control of T7 promoter in Escherichia coli. The recombinant proteins were purified to achieve homogeneity using Ni-agarose column chromatography. The molecular mass of the purified DszA and DszB were determined to be 51.9 and 39.2 kDa respectively by using SDS-polyacrylamide gel electrophoresis. DszA showed a K m of 0.14±0.005 mM and a maximal velocity of 0.004±0.0004 mM/min. DszB showed a wide substrate range in a way that all aromatic sulfonates compounds acted as its substrate; as it seemed the active site was suitable for the sulfonated aromatic rings. The K m and V max values of DszB were calculated to be 1.81±0.02 mM and 6.55 ± 0.005 µM/min respectively using 4-Amino-3-hydroxy-naphthalene-sulfonic acid as a substrate

Developing a circularly permuted variant of Renilla luciferase as a bioluminescent sensor for measuring Caspase-9 activity in the cell-free and cell-based systems

Biosensors and whole cell biosensors consisting of biological molecules and living cells can sense a special stimulus on a living system and convert it to a measurable signal. A major group of them are the bioluminescent sensors derived from luciferases. This type of biosensors has a broad application in molecular biology and imaging systems. In this project, a luciferase-based biosensor for detecting and measuring caspase-9 activity is designed and constructed using the circular permutation strategy. The spectroscopic method results reveal changes in the biosensor structure. Additionally, its activity is examined in a cell-free coupled assay system. Afterward, the biosensor is utilized for measuring the cellular caspase-9 activity upon apoptosis induction in a cancer cell line. In following the gene of biosensor is sub-cloned into a eukaryotic vector and transfected to HEK293T cell line and then its activity is measured upon apoptosis induction in the presence and absence of a caspase-9 inhibitor. The obtained results show that the designed biosensor detects the caspase-9 activity in the cell-free and cell-based systems

Developing a circularly permuted variant of Renilla luciferase as a bioluminescent sensor for measuring Caspase-9 activity in the cell-free and cell-based systems

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