Abstract:Inosine-5′-monophosphate dehydrogenase (IMPDH) plays key roles in purine nucleotide metabolism and cell proliferation. Although IMPDH is a widely studied therapeutic target, there is limited information about its physiological regulation. Using Ashbya gossypii as a model, we describe the molecular mechanism and the structural basis for the allosteric regulation of IMPDH by guanine nucleotides. We report that GTP and GDP bind to the regulatory Bateman domain, inducing octamers with compromised catalytic activit… Show more
“…5), pointing to an important physiological role of the IMPDH conformational switch within cells.Figure 2The conformational switch of AgIMPDH. ( A ) High-resolution structures of AgIMPDH in complex with ATP (left panel in blue cartoons) and GDP (right panel in green cartoons 5 ). ATP and GDP molecules are shown in orange and blue sticks, respectively.…”
Inosine-5′-monophosphate dehydrogenase (IMPDH) is an essential enzyme for nucleotide metabolism and cell proliferation. Despite IMPDH is the target of drugs with antiviral, immunosuppressive and antitumor activities, its physiological mechanisms of regulation remain largely unknown. Using the enzyme from the industrial fungus Ashbya gossypii, we demonstrate that the binding of adenine and guanine nucleotides to the canonical nucleotide binding sites of the regulatory Bateman domain induces different enzyme conformations with significantly distinct catalytic activities. Thereby, the comparison of their high-resolution structures defines the mechanistic and structural details of a nucleotide-controlled conformational switch that allosterically modulates the catalytic activity of eukaryotic IMPDHs. Remarkably, retinopathy-associated mutations lie within the mechanical hinges of the conformational change, highlighting its physiological relevance. Our results expand the mechanistic repertoire of Bateman domains and pave the road to new approaches targeting IMPDHs.
“…5), pointing to an important physiological role of the IMPDH conformational switch within cells.Figure 2The conformational switch of AgIMPDH. ( A ) High-resolution structures of AgIMPDH in complex with ATP (left panel in blue cartoons) and GDP (right panel in green cartoons 5 ). ATP and GDP molecules are shown in orange and blue sticks, respectively.…”
Inosine-5′-monophosphate dehydrogenase (IMPDH) is an essential enzyme for nucleotide metabolism and cell proliferation. Despite IMPDH is the target of drugs with antiviral, immunosuppressive and antitumor activities, its physiological mechanisms of regulation remain largely unknown. Using the enzyme from the industrial fungus Ashbya gossypii, we demonstrate that the binding of adenine and guanine nucleotides to the canonical nucleotide binding sites of the regulatory Bateman domain induces different enzyme conformations with significantly distinct catalytic activities. Thereby, the comparison of their high-resolution structures defines the mechanistic and structural details of a nucleotide-controlled conformational switch that allosterically modulates the catalytic activity of eukaryotic IMPDHs. Remarkably, retinopathy-associated mutations lie within the mechanical hinges of the conformational change, highlighting its physiological relevance. Our results expand the mechanistic repertoire of Bateman domains and pave the road to new approaches targeting IMPDHs.
“…Depletion of guanine allows Bateman domain extension, and transient assembly into enzymatically active filaments. However, upon restoration of guanine levels, these active filaments disassemble into compressed free octamers, mirroring the known feedback inhibition behavior of non-filament forming IMPDH homologues (Buey, Ledesma-Amaro, Velázquez-Campoy, et al 2015; Buey et al 2017; Fernández-Justel et al 2019).…”
Section: Discussionmentioning
confidence: 82%
“…Sites 1 and 2 are canonical cystathionine beta synthase motifs that bind either ATP/ADP or GTP/GDP, and are conserved among IMPDH homologues (Scott et al 2004; Ignoul & Eggermont 2005; Baykov et al 2011; Ereño-Orbea et al 2013). Site 3 is a non-canonical site located at the interface between domains that binds only GTP/GDP (Buey, Ledesma-Amaro, Velázquez-Campoy, et al 2015).…”
Section: Introductionmentioning
confidence: 99%
“…GTP/GDP binding induces a compressed conformation by changing the relative orientation of the two domains. This brings the active sites of opposing tetramers tightly together, forming an interdigitated pseudo beta-barrel that prevents reopening and product release and inhibits IMPDH activity by a mechanism described as a “conformational switch” between extended and compressed states (Buey, Ledesma-Amaro, Velázquez-Campoy, et al 2015; Buey et al 2017; Fernández-Justel et al 2019). Because inhibition requires interactions between two opposing tetramers, this switch is functionally relevant in only the octameric state.…”
8 Inosine monophosphate dehydrogenase (IMPDH) mediates the first committed step in guanine 9 nucleotide biosynthesis and plays important roles in cellular proliferation and the immune response. The 10 enzyme is heavily regulated to maintain balance between guanine and adenine nucleotide pools. IMPDH 11 reversibly polymerizes in cells and tissues in response to changes in metabolic demand, providing an 12 additional layer of regulatory control associated with increased flux through the guanine synthesis 13 pathway. Here, we report a series of human IMPDH2 cryo-EM structures in active and inactive 14 conformations, and show that the filament resists inhibition by guanine nucleotides. The structures define 15 the mechanism of filament assembly, and reveal how assembly interactions tune the response to guanine 16 inhibition. Filament-dependent allosteric regulation of IMPDH2 makes the enzyme less sensitive to 17 feedback inhibition, explaining why assembly occurs under physiological conditions, like stem cell 18 proliferation and T-cell activation, that require expansion of guanine nucleotide pools.
“…In eukaryotes, it has been reported that GDP/GTP binds to the CBS domain of IMPDH and allosterically inhibits its enzymatic activity. However, this is not observed in prokaryotes (Buey et al, , ). Furthermore, it has been reported that single‐stranded nucleic acids bind to this subdomain in human IMPDH (Mclean et al, ).…”
In bacteria, guanosine (penta)tetra‐phosphate ([p]ppGpp) is essential for controlling intracellular metabolism that is needed to adapt to environmental changes, such as amino acid starvation. The (p)ppGpp0 strain of Bacillus subtilis, which lacks (p)ppGpp synthetase, is unable to form colonies on minimal medium. Here, we found suppressor mutations in the (p)ppGpp0 strain, in the purine nucleotide biosynthesis genes, prs, purF and rpoB/C, which encode RNA polymerase core enzymes. In comparing our work with prior studies of ppGpp0 suppressors, we discovered that methionine addition masks the suppression on minimal medium, especially of rpoB/C mutations. Furthermore, methionine addition increases intracellular GTP in rpoB suppressor and this effect is decreased by inhibiting GTP biosynthesis, indicating that methionine addition activated GTP biosynthesis and inhibited growth under amino acid starvation conditions in (p)ppGpp0 backgrounds. Furthermore, we propose that the increase in intracellular GTP levels induced by methionine is due to methionine derivatives that increase the activity of the de novo GTP biosynthesis enzyme, GuaB. Our study sheds light on the potential relationship between GTP homeostasis and methionine metabolism, which may be the key to adapting to environmental changes.
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