GAPDH delivers heme to soluble guanylyl cyclase. J Biol Chem 2020 Jun 12;295(24):8145-8154
Date
05/03/2020Pubmed ID
32358060Pubmed Central ID
PMC7294094DOI
10.1074/jbc.RA120.013802Scopus ID
2-s2.0-85086495703 (requires institutional sign-in at Scopus site) 39 CitationsAbstract
Soluble guanylyl cyclase (sGC) is a key component of NO-cGMP signaling in mammals. Although heme must bind in the sGC β1 subunit (sGCβ) for sGC to function, how heme is delivered to sGCβ remains unknown. Given that GAPDH displays properties of a heme chaperone for inducible NO synthase, here we investigated whether heme delivery to apo-sGCβ involves GAPDH. We utilized an sGCβ reporter construct, tetra-Cys sGCβ, whose heme insertion can be followed by fluorescence quenching in live cells, assessed how lowering cell GAPDH expression impacts heme delivery, and examined whether expressing WT GAPDH or a GAPDH variant defective in heme binding recovers heme delivery. We also studied interaction between GAPDH and sGCβ in cells and their complex formation and potential heme transfer using purified proteins. We found that heme delivery to apo-sGCβ correlates with cellular GAPDH expression levels and depends on the ability of GAPDH to bind intracellular heme, that apo-sGCβ associates with GAPDH in cells and dissociates when heme binds sGCβ, and that the purified GAPDH-heme complex binds to apo-sGCβ and transfers its heme to sGCβ. On the basis of these results, we propose a model where GAPDH obtains mitochondrial heme and then forms a complex with apo-sGCβ to accomplish heme delivery to sGCβ. Our findings illuminate a critical step in sGC maturation and uncover an additional mechanism that regulates its activity in health and disease.
Author List
Dai Y, Sweeny EA, Schlanger S, Ghosh A, Stuehr DJAuthor
Elizabeth Sweeny PhD Assistant Professor in the Biochemistry department at Medical College of WisconsinMESH terms used to index this publication - Major topics in bold
AnimalsApoproteins
Glyceraldehyde-3-Phosphate Dehydrogenase (Phosphorylating)
HEK293 Cells
Heme
Humans
Kinetics
Mitochondria
Models, Biological
Protein Binding
Protein Multimerization
Rats
Soluble Guanylyl Cyclase
