Oxaloacetate Production via Carboxylations in <i>Crithidia fasciculata</i> Preparations

Abstract

SYNOPSIS. Fractions containing soluble enzymes from Crithidia fasciculata had an ADP‐linked phosphoenolpyruvate (PEP) carboxykinase. The enzyme produced ATP and oxaloacetate (OAA) from PEP, ADP and HCO − 3 . OAA was determined as the endproduct of reactions by forming the 2,4‐dinitrophenylhydrazone derivative; the hydrazone was identified by thin‐layer chromatography. Approximate Michaelis constants (PEP, Mg, HCO − 3 , ADP) were determined spectrophotometrically by linking OAA production to malic dehydrogenase. The PEP carboxykinase did not utilize GDP, UDP or IDP as cofactors; the metal requirement was also satisfied by Mn. The enzyme was inhibited by the biotin antagonists avidin and desthiobiotin. A pyruvate carboxylase was also present in the preparations, generating OAA from pyruvate and ATP. The role of both enzymes in OAA production and subsequent production of succinate is discussed with regard to C. fasciculata and other trypanosomatids.