dGPredictor: Automated fragmentation method for metabolic reaction free energy prediction and de novo pathway design

Group contribution (GC) methods are conventionally used in thermodynamics analysis of metabolic pathways to estimate the standard Gibbs free energy change (ΔrG′o) of enzymatic reactions from limited experimental measurements. However, these methods are limited by their dependence on manually curated groups and inability to capture stereochemical information, leading to low reaction coverage. Herein, we introduce an automated molecular fingerprint-based thermodynamic analysis tool called dGPredictor that enables the consideration of stereochemistry within metabolite structures and thus increases reaction coverage. dGPredictor has a higher prediction accuracy as compared to existing GC methods and can capture free energy changes for isomerase and transferase reactions, which exhibit no overall group changes. We also demonstrate dGPredictor’s ability to predict the Gibbs free energy change for novel reactions and seamless integration within de novo metabolic pathway design tools such as novoStoic. This enables performing a thermodynamic analysis for synthetic pathways, thus safeguarding against the inclusion of reaction steps with infeasible directionalities. To facilitate easy access to dGPredictor, we developed a graphical user interface to predict the standard Gibbs free energy change for reactions at various pH and ionic strengths. The tool allows customized user input of known metabolites as KEGG IDs and novel metabolites as InChI strings (https://github.com/maranasgroup/dGPredictor).Author summaryThe genome-scale metabolic networks consist of a large number of biochemical reactions interconnected in a complex system. The standard Gibbs free energy change is commonly used to check for the feasibility of enzyme-catalyzed reactions as thermodynamics plays a crucial role in pathway design for biochemical synthesis. The group contribution methods using expert-defined functional groups have been extensively used for estimating standard Gibbs free energy change with limited experimental measurements. However, current methods using functional groups have major issues that limit its ability to cover all the metabolites and reactions as well as the inability to consider stereochemistry leads to erroneous estimation of free energy that undergoes only stereochemical change such as isomerases. Here, we introduce a molecular fingerprint-based thermodynamic tool dGPredictor that enables stereochemistry in metabolites and thus improves the reaction coverage with higher prediction accuracy compared to current GC methods. It also allows the ability to predict free energy change for novel reactions which can aid the de novo metabolic pathway design tool to ensure the reaction feasibility. We apply and test our method on reactions in the KEGG database and isobutanol synthesis pathway. In addition, we provide an open-source user-friendly web interface to facilitate easy access for standard Gibbs free energy change of reactions at different physiological states.

Biotransformation potential of phytosterols under anoxic and anaerobic conditions

The biotransformation potential of three phytosterols (campesterol, stigmasterol and β-sitosterol) under denitrifying, sulfate-reducing and fermentative/methanogenic conditions was assessed. Using a group contribution method, the standard Gibbs free energy of phytosterols was calculated and used to perform theoretical energetic calculations. The oxidation of phytosterols under aerobic, nitrate-reducing, sulfate-reducing and methanogenic conditions was determined to be energetically feasible. However, using semi-continuously fed cultures maintained at 20–22 °C over 16 weekly feeding cycles (112 days; retention time, 21 days), phytosterol removal was observed under nitrate-reducing and sulfate-reducing conditions, but not under fermentative/methanogenic conditions. Under sulfate-reducing conditions, stigmast-4-en-3-one was identified as an intermediate of phytosterol biotransformation, a reaction more likely carried out by dehydrogenases/isomerases, previously reported to act on cholesterol under both oxic and anoxic (denitrifying) conditions. Further study of the biotransformation of phytosterols under anoxic/anaerobic conditions is necessary to delineate the factors and conditions leading to enhanced phytosterol biodegradation and the development of effective biological treatment systems for the removal of phytosterols from pulp and paper wastewaters and other phytosterol-bearing waste streams.

CMC Determination and Thermodynamic Micellisation Of NPE Surfactant In Aqueous And CH3OH – H2O Solvents

The critical micelle concentration (CMC) of nonylphenolethoxylate (NPE) surfactant has been determined by measuring the surface tension as a function of the molar concentration of the surfactant in aqueous and binary mixture of water + methanol solutions at a temperature range from 20?C to 35?C. The interfacial parameters ?max, Amin, ?cmc and ?G?ads were calculated. The results indicate that the CMC increases as the temperature increases and that the addition of methanol the CMC decreases. The thermodynamic parameters such as standard Gibbs free energy (?G?), enthalpy (?H?), and entropy (?S?) of micellization were estimated using the change of CMC with temperature. The enthalpy – entropy compensation behavior of the surfactant was evaluated and a good linearity in the compensation plot has been observed.