COMPLEMENTATION OF AN ESCHERICHIA-COLI ADHE MUTANT BY THE ENTAMOEBA-HISTOLYTICA EHADH2 GENE PROVIDES A METHOD FOR THE IDENTIFICATION OF NEWANTIAMEBIC DRUGS

The pathogenic protozoan parasite Entamocba histolytica, the cause of amebic dysentery and amebic liver abscess, is an obligate anaerobe, an d derives energy from the fermentation of glucose to ethanol with pyru vate and acetyl coenzyme A as intermediates. We have isolated EhADH2, a key enzyme in this pathway, that is a NAD(+)-and Fe2+-dependent bifu nctional enzyme with acetaldehyde dehydrogenase and alcohol dehydrogen ase activities, EhADH2 is tile only known eukaryotic member of a newly defined family of prokaryotic multifunctional enzymes, which includes the Escherichia coli AdhE enzyme, an enzyme required for anaerobic gr owth of E. coli. Because of the critical role of EhADH2 in the amebic fermentation pathway and the lack of known eukaryotic homologues of th e EhADH2 enzyme, EhADH2 represents a potential target for antiamebic c hemotherapy. However, screening of compounds for antiamebic activity I s hampered by the cost of large scale growth of Ent. histolytica, and difficulties in quantitating drug efficacy in vitro. To approach this problem, we expressed the EhADH2 gene in a mutant strain of E. coti ca rrying a deletion of the adhE gent, Expression of EhADH2 restored the ability of the mutant E. coli strain to grow under anaerobic condition s, By screening compounds for the ability to inhibit the anaerobic gro wth of the E. coli/EhADH2 strain, we have developed a rapid assay for identifying compounds with, anti-EhADH2 activity. Using bacteria to by pass the need for parasite culture in the initial screening process fo r anti-parasitic agents could greatly simplify and reduce the cost of identifying new therapeutic agents effective against parasitic disease s.