δ1-Pyrroline-5-carboxylate reductase as a new target for therapeutics: inhibition of the enzyme from Streptococcus pyogenes and effects in vivo

Abstract

Compounds able to interfere with amino acid biosynthesis have the potential to inhibit cell growth. In both prokaryotic and eukaryotic microorganisms, unless an ornithine cyclodeaminase is present, the activity of δ1-pyrroline-5-carboxylate (P5C) reductase is mandatory to proline production, and the enzyme inhibition should result in amino acid starvation, blocking in turn protein synthesis. The ability of some substituted derivatives of aminomethylenebisphosphonic acid and its analogues to interfere with the activity of the enzyme from the human pathogen Streptococcus pyogenes was investigated. Several compounds were able to suppress activity in the micromolar range of concentrations, with a mechanism of uncompetitive type with respect to the substrate P5C and non-competitive with respect to the electron donor NAD(P)H. The actual occurrence of enzyme inhibition in vivo was supported by the effects of the most active derivatives upon bacterial growth and free amino acid content.

Coenzyme preference of Streptococcus pyogenes δ1-pyrroline-5-carboxylate reductase: evidence supporting NADPH as the physiological electron donor

“…P5CR activity was measured by following the P5C-dependent oxidation of NAD(P)H (Forlani et al 2011). For kinetic evaluations, curve fitting by non-linear regression analysis was performed with Prism 5 (GraphPad Software).…”

“…Transformation, induction and affinity purification of the enzyme were as described previously (Forlani et al 2011). Protein concentration was determined by the method of Bradford (1976), using bovine serum albumin as the standard.…”

“…Some phosphonate inhibitors were indeed found to exert both phytotoxic (Forlani et al 2007(Forlani et al , 2008 and antibiotic (Forlani et al 2011) effects. Here we report the results of a thorough biochemical characterization of P5CR from S. pyogenes, shedding more light on its substrate preference and reaction mechanism.…”

Physiologic Activity of Bisphosphonates – Recent Advances

“…Other enzymes have also been considered as targets for specific antibacterial agents, namely thymidyltransferase [186] and δ 1 -pyrroline-5-carboxylate reductase as an inhibitor of Streptococcus pyogenes , [187] which is the cause of many important human diseases ranging from mild superficial skin infections to life-threatening systemic infections. Additionally, glutamine synthetase could also be a target of a selective anti-tuberculosis agent for use against infections of the musculoskeletal system [188].…”

Synthetic Procedures Leading towards Aminobisphosphonates

“…Because this compound was previously prepared by acid hydrolysis of N-benzhydrylaminomethylenebisphosphonic acid [83,100], it may be that it is formed upon hydrolysis of N-aryl derivatives. The three-component reaction was applied to synthesize a large series of physiologically active compounds, in some cases of very complex chemical structures, including bone antiresorptive drug candidates [75][76][77][78][79][80][81][82]; bone imaging [83,84], antiprotozoal [85][86][87][88], antibacterial [72,[89][90][91][92], anti-HIV [93] and anti-inflammatory [94] agents; herbicides [95][96][97]; and complex ones for various metals [10,98].…”