title: Rapid determination of tricarboxylic acid cycle enzyme activities in biological samples.
abstract: BACKGROUND
  In the last ten years, deficiencies in tricarboxylic acid cycle (TCAC) enzymes have been shown to cause a wide spectrum of human diseases, including malignancies and neurological and cardiac diseases. A prerequisite to the identification of disease-causing TCAC enzyme deficiencies is the availability of effective enzyme assays.
  
  
  RESULTS
  We developed three assays that measure the full set of TCAC enzymes. One assay relies on the sequential addition of reagents to measure succinyl-CoA ligase activity, followed by succinate dehydrogenase, fumarase and, finally, malate dehydrogenase. Another assay measures the activity of alpha-ketoglutarate dehydrogenase followed by aconitase and isocitrate dehydrogenase. The remaining assay measures citrate synthase activity using a standard procedure. We used these assays successfully on extracts of small numbers of human cells displaying various severe or partial TCAC deficiencies and on frozen heart homogenates from heterozygous mice harboring an SDHB gene deletion.
  
  
  CONCLUSIONS
  This set of assays is rapid and simple to use and can immediately detect even partial defects, as the activity of each enzyme can be readily compared with one or more other activities measured in the same sample.
authors.affiliation: Inserm, U676, Paris, 75019 France.
authors.forename: Sergio
authors.initials: S
authors.lastname: Goncalves
authors.forename: Vincent
authors.initials: V
authors.lastname: Paupe
authors.forename: Emmanuel P
authors.initials: EP
authors.lastname: Dassa
authors.forename: Jean-Jacques
authors.initials: JJ
authors.lastname: Brière
authors.forename: Judith
authors.initials: J
authors.lastname: Favier
authors.forename: Anne-Paule
authors.initials: AP
authors.lastname: Gimenez-Roqueplo
authors.forename: Paule
authors.initials: P
authors.lastname: Bénit
authors.forename: Pierre
authors.initials: P
authors.lastname: Rustin
mesh_terms: D000154:Aconitate Hydratase / Q000378:metabolism; D000818:Animals; D002950:Citrate (si)-Synthase / Q000378:metabolism; D002952:Citric Acid Cycle; D057075:Enzyme Assays / Q000379:methods*; D004798:Enzymes / Q000378:metabolism; D005649:Fumarate Hydratase / Q000378:metabolism; D007521:Isocitrate Dehydrogenase / Q000378:metabolism; D007655:Ketoglutarate Dehydrogenase Complex / Q000378:metabolism; D008291:Malate Dehydrogenase / Q000378:metabolism; D051379:Mice; D009206:Myocardium / Q000201:enzymology; D013385:Succinate Dehydrogenase / Q000378:metabolism; D013389:Succinate-CoA Ligases / Q000378:metabolism
chemical_list: D004798:Enzymes; D008291:Malate Dehydrogenase; D007521:Isocitrate Dehydrogenase; D007655:Ketoglutarate Dehydrogenase Complex; D013385:Succinate Dehydrogenase; D002950:Citrate (si)-Synthase; D005649:Fumarate Hydratase; D000154:Aconitate Hydratase; D013389:Succinate-CoA Ligases