A SINGLE BASE-PAIR CHANGE (ATG-]ATC) NULLIFIES THE ACTIVITY OF CYTOSOLIC FUMARASE IN SACCHAROMYCES-CEREVISIAE

A respiratory defective pet mutant (W303 del FUM1) of Saccharomyces ce revisiae deficient in fumarase was transformed with the plasmid constr uct pG5/ST7. This plasmid contains the entire FUM1 gene with only a si ngle base pair change (ATG --> ATC) confined to the putative second in -frame translation initiation codon. While transformation of the fumar ase deficient mutant with pG5/ST7 resulted in an elevation of fumarase activity in the mitochondria of the transformed strain, fumarase acti vity in the cytosol remained negligible. This result indicated that th e cytosolic fumarase isoenzyme is synthesized exclusively from the sec ond translation initiation codon of FUM1. Results of RACE-PCR of the 5 ' ends of FUM1 transcripts confirmed that the FUM1 gene synthesizes tw o distinct species of transcripts. These data provide strong evidence for the synthesis and targeting of two fumarase isomers. (C) 1995 Acad emic Press, Inc.