A metabolic control analysis approach to introduce the study of systems in biochemistry: the glycolytic pathway in the red blood cell

ANGELANI C; CARABIAS P; CRUZ K; DELFINO J; DE SAUTU M; ESPELT MV; FERREIRA GOMES M; GOMES G; MANGIALAVORI I; MANZI M; PIGNATARO F; SAFFIOTI N; SALVATIERRA D; SANTOS J; SCHWARZBAUM P

BIOCHEMISTRY AND MOLECULAR BIOLOGY EDUCATION

JOHN WILEY & SONS INC

Lugar: Nueva York; Año: 2018

1470-8175

Review2AbstractMetabolic Control Analysis (MCA) is a promising approach in biochemistry aimed at understandingprocesses in a quantitative fashion. Here the contribution of enzymes and transporters to the control of agiven pathway flux and metabolite concentrations is determined and expressed quantitatively by means ofnumerical coefficients. Metabolic flux can be influenced by a wide variety of modulators acting on one ormore metabolic steps along the pathway.We describe a laboratory exercise to study metabolic regulation of human erythrocytes (RBCs).Within the framework of MCA, students use these cells to determine the sensitivity of the glycolytic flux totwo inhibitors (iodoacetic acid: IA, and iodoacetamide: IAA) known to act on the enzyme glyceraldehyde-3-phosphate-dehydrogenase. Glycolytic flux was estimated by determining the concentration of extracellularlactate, the end product of RBC glycolysis. A low-cost colorimetric assay was implemented, that takesadvantage of the straightforward quantification of the absorbance signal from the photographic image of themulti-well plate taken with a standard digital camera.Students estimate flux response coefficients for each inhibitor by fitting an empirical function to theexperimental data, followed by analytical derivation of this function. IA and IAA exhibit qualitativelydifferent patterns, which are thoroughly analyzed in terms of the physicochemical properties influencingtheir action on the target enzyme. IA causes highest glycolytic flux inhibition at lower concentration thanIAA. This work illustrates the feasibility of using the MCA approach to study key variables of a simplemetabolic system, in the context of an upper level biochemistry course