title: Thiamin auxotrophy in yeast through altered cofactor dependence of the enzyme acetohydroxyacid synthase.
abstract: The THI1 gene of Saccharomyces cerevisiae has been identified and found to be allelic with the previously characterized gene ILV2 that encodes acetohydroxyacid synthase (AHAS). This enzyme catalyses the first step in the parallel biosyntheses of the branched-chain amino acids isoleucine and valine, using thiamin pyrophosphate (TPP) as a cofactor. The ilv2-thi1 allele encodes a functional AHAS enzyme with an altered dependence for the cofactor TPP resulting in the thiamin auxotrophic phenotype. Nucleotide sequence analysis and site-directed mutagenesis revealed that the thi1 mutation is a single base substitution which causes the conserved amino acid substitution D176E in the AHAS protein. This study therefore implicates aspartate 176 as another amino acid residue important either for the efficient binding of TPP by AHAS or for the functional stability of the holoenzyme.
authors.affiliation: Department of Genetics, University of Leicester, University Road, Leicester LE1 7RH, UK1.
authors.forename: Kerry L
authors.initials: KL
authors.lastname: Byrne
authors.affiliation: Department of Genetics, University of Leicester, University Road, Leicester LE1 7RH, UK1.
authors.forename: Peter A
authors.initials: PA
authors.lastname: Meacock
mesh_terms: D000095:Acetolactate Synthase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D000483:Alleles; D000595:Amino Acid Sequence; D001483:Base Sequence; D020134:Catalytic Domain / Q000235:genetics; D003001:Cloning, Molecular; D004271:DNA, Fungal / Q000235:genetics; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D005800:Genes, Fungal; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010641:Phenotype; D010957:Plasmids / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D013831:Thiamine / Q000378:metabolism*; D013835:Thiamine Pyrophosphate / Q000378:metabolism; D014157:Transcription Factors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: D004271:DNA, Fungal; D004268:DNA-Binding Proteins; D014157:Transcription Factors; D000095:Acetolactate Synthase; D013835:Thiamine Pyrophosphate; D013831:Thiamine