Structure‐Based Insights into Control of Heme Biosynthesis

Abstract

Protein assembly drives many aspects of cellular and mitochondrial physiology. We are interested in uncovering and characterizing the protein interactions that support proper heme biosynthesis. Heme is a critical biomolecule that carries out several functions in nearly all life forms, including humans where its most widely‐known role is mediating oxygen transport in blood. It is imperative that heme production is tightly controlled as alterations in cellular heme levels can have drastic consequences for human health. The first and rate‐limiting enzyme controlling heme biosynthesis is aminolevulinic acid synthase (ALAS). ALAS is conserved in α‐proteobacteria and non‐plant eukaryotes, there are two isoforms in vertebrates, with the mitochondrial protein ALAS2 being responsible for heme synthesis during erythropoiesis. There are two separate diseases that may result from one of over 50 mutations in the ALAS2 gene. Mutations decreasing optimal activity lead to the disease X‐linked sideroblastic anemia (XLSA), whereas other mutations causing hyperactivity underlie X‐linked protoporphyria (XLPP). Importantly, several disease‐causing mutations are located in a eukaryote‐specific C‐terminal extension, a region absent from bacterial ALAS enzymes. Certain mutations also result in an inability of ALAS2 to assemble with other proteins, including proteins involved in separate metabolic pathways. However, we lack an understanding of how these mutations lead to a change in ALAS2 structure, and therefore, function. We seek to understand how ALAS interacts with accessory proteins as well as organism‐specific differences in assembly that may alter regulation of heme production. Therefore, we use X‐ray crystallography combined with in vitro activity assays of various eukaryotic ALAS enzymes to parse apart the role of this key regulatory region in eukaryote ALAS function. Our work is beginning to reveal key structure‐function relationships between the orientation and molecular contacts mediated by the C‐terminus and overall ALAS enzyme function, thus controlling heme biosynthesis.Support or Funding InformationJane Coffin Childs Memorial Fund for Medical Research Award and a Burroughs Wellcome Postdoctoral Enrichment Program Fellowship Award 1015092 (B.L.B); NIH Ruth L. Kirschstein National Research Service Award (F32DK095726) (J.R.K.); National Institutes of Health Grant (R01 DK115558) (T.A.B.), and the Howard Hughes Medical Institute. X‐ray data collected at NE‐CAT (P41 GM103403).