PT  - JOURNAL ARTICLE
AU  - Schmitz, Jonathan M.
AU  - Wolters, John F.
AU  - Murray, Nathan H.
AU  - Guerra, Rachel M.
AU  - Bingman, Craig A.
AU  - Hittinger, Chris Todd
AU  - Pagliarini, David J.
TI  - Aim18p and Aim46p are CHI-domain-containing mitochondrial hemoproteins in <em>Saccharomyces cerevisiae</em>
AID  - 10.1101/2022.11.15.516536
DP  - 2022 Jan 01
TA  - bioRxiv
PG  - 2022.11.15.516536
4099  - http://biorxiv.org/content/early/2022/11/15/2022.11.15.516536.short
4100  - http://biorxiv.org/content/early/2022/11/15/2022.11.15.516536.full
AB  - Chalcone isomerases (CHIs) have well-established roles in the biosynthesis of plant flavonoid metabolites. Saccharomyces cerevisiae possesses two predicted CHI-like proteins, Aim18p (encoded by YHR198C) and Aim46p (YHR199C), but it lacks other enzymes of the flavonoid pathway, suggesting that Aim18p and Aim46p employ the CHI fold for distinct purposes. Here, we demonstrate that Aim18p and Aim46p reside on the mitochondrial inner membrane and adopt CHI folds, but they lack select active site residues and possess an extra fungal-specific loop. Consistent with these differences, Aim18p and Aim46p lack chalcone isomerase activity and also the fatty acid-binding capabilities of other CHI-like proteins, but instead bind heme. We further show that diverse fungal homologs also bind heme and that Aim18p and Aim46p possess structural homology to a bacterial hemoprotein. Collectively, our work reveals a distinct function and cellular localization for two CHI-like proteins, introduces a new variation of a hemoprotein fold, and suggests that ancestral CHI-like proteins were hemoproteins.Competing Interest StatementThe authors have declared no competing interest.CHIchalcone isomeraseCHILchalone isomerase likeFAPfatty acid binding proteinDNAdeoxyribonucleic acidHMG-CoAβ-Hydroxy β-methylglutaryl-CoANdxxN-terminal deletion of xx amino acidsORFopen reading frameSCsynthetic completeUra-uracil drop-out mediaGPDglyceraldehyde-3-phosphate dehydrogenaseSSsalmon sperm DNAPEGpolyethylene glycolFLAGimmunoprecipitation epitope consisting of Asp-Tyr-Lys-Asp-Asp-Asp-Asp-Lysbpbase pairsSDMsite-directed mutagenesisBMEβ-mercaptoethanolTCAtrichloroacetic acidLDSlithium dodecyl sulfateSDS-PAGEsodium dodecyl sulfate–polyacrylamide gel electrophoresisPVDFpolyvinylidene fluorideNFDMnon-fat dry milkTBSTtris buffered saline plus 0.05% tween-20 (v/v)(v/v)percent volume per 100 mL volume(w/v)percent weight per 100 mL volumeGAPDHglyceraldehyde-3-phosphate dehydrogenaseCYTBcytochrome bCITcitrate synthaseTOMtranslocase of the outer membraneTIMtranslocase of the inner membraneSECsize-exclusion chromatographyTCEPtris(2-carboxyethyl)phosphineDTTdithiothreitolDSFdifferential scanning fluorimetryUV-Visultraviolet–visible spectroscopyCoQcoenzyme Q or ubiquinoneDMQdemethoxyubiquinonePPHBpolyprenyl-4-hydroxybenzoatePDBProtein Data Bank