Immobilization of arginase and its application in an enzymatic chromatographic column: Thermodynamic studies of nor-NOHA/arginase binding and role of the reactive histidine residue

Experimental studies of OH° radical/pressure dependence of arginase activity using a molecular chromatography approach

“…These results indicated that the IMER could be used to on line screen for new inhibitors and the enzyme immobilization on the chromatographic support did not alter its biological properties. To evaluate the IMER to IMER reproducibility, three IMERs were prepared under identical conditions [16]. The mobile phase A was 0.1 mM Tris-HCl buffer pH = 7.4-10 mM MnCl 2 , the IMER was maintained equal to 25 • C at a flow-rate of 0.5 mL/min.…”

“…The preparation of the immobilized enzyme reactor (IMER) using a monolithic support inside a column (25 mm × 4.6 mm i.d.) was given in [16]. The mobile phase A was described below and depended on the developed application.…”

“…The ex vivo characterization of drug candidates on isolated target enzymes, unfortunately, is often associated with long, labor-intensive assays and a large amount of disposable expensive material. Our group recently developed a novel immobilized arginase reactor for the binding mechanism study of a series of arginase inhibitors with the enzyme and the magnesium effect on this association process [16,17]. This paper describes a novel procedure for studying both the direct role of OH • radical formation and pressure on the arginase activity.…”

Biochromatographic framework for analyzing magnesium chloride salt dependence on nor-NOHA binding to arginase enzyme

“…Recently, our group demonstrated that arginase I inhibition reduces endothelial dysfunction and blood pressure rising in spontaneously hypertensive rats [1,2]. A novel chromatographic support was as well developed to determine and quantify the forces driving association between N -hydroxy-norl-arginine (nor-NOHA) which is a very good arginase inhibitor [3]. The energetic of binding of the inhibitor to the enzyme as both a function of temperature and pH was studied using this novel biochromatographic approach [3].…”

“…A novel chromatographic support was as well developed to determine and quantify the forces driving association between N -hydroxy-norl-arginine (nor-NOHA) which is a very good arginase inhibitor [3]. The energetic of binding of the inhibitor to the enzyme as both a function of temperature and pH was studied using this novel biochromatographic approach [3]. Magnesium cation Mg 2+ is an important factor in the physiology of cardiovascular apparatus and the pathogenesis of cardiovascular diseases.…”