Intersubunit Coupling Enables Fast CO 2 -Fixation by Reductive Carboxylases
Not Available
37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
Engineering topics
Vöhringer-Martinez, Esteban
Publications and source records attributed to Vöhringer-Martinez, Esteban.
Long-range structural defects by pathogenic mutations in most severe glucose-6-phosphate dehydrogenase deficiency
Glucose-6-phosphate dehydrogenase (G6PD) deficiency is the most common blood disorder, presenting multiple symptoms, including hemolytic anemia. It affects 400 million people worldwide, with more than 160 single mutations reported in G6PD. The most severe mutations (about 70) are classified as class I, leading to more than 90% loss of activity of the wild-type G6PD. The crystal structure of G6PD reveals these mutations are located away from the active site, concentrating around the noncatalytic NADP + -binding site and the dimer interface. However, the molecular mechanisms of class I mutant dysfunction have remained elusive, hindering the development of efficient therapies. To resolve this, we performed integral structural characterization of five G6PD mutants, including four class I mutants, associated with the noncatalytic NADP + and dimerization, using crystallography, small-angle X-ray scattering (SAXS), cryogenic electron microscopy (cryo-EM), and biophysical analyses. Comparisons with the structure and properties of the wild-type enzyme, together with molecular dynamics simulations, bring forward a universal mechanism for this severe G6PD deficiency due to the class I mutations. We highlight the role of the noncatalytic NADP + -binding site that is crucial for stabilization and ordering two β-strands in the dimer interface, which together communicate these distant structural aberrations to the active site through a network of additional interactions. This understanding elucidates potential paths for drug development targeting G6PD deficiency.
59 BASIC BIOLOGICAL SCIENCES↗
Structural adaptation of oxygen tolerance in 4-hydroxybutyrl-CoA dehydratase, a key enzyme of archaeal carbon fixation
Abstract Autotrophic microorganisms that convert inorganic carbon into organic matter were key players in the evolution of life on Earth. As the early atmosphere became oxygenated, microorganisms needed to develop mechanisms for oxygen protection, especially those relying on enzymes containing oxygen-sensitive metal clusters (e.g., Fe-S). Here we investigated how 4-hydroxybutyryl-CoA dehydratase (4HBD) - the key enzyme of the 3-hydroxypropionate/4-hydroxybutyrate (HP/HB) cycle for CO 2 -fixation - adapted as conditions shifted from anoxic to oxic. 4HBD is found in both anaerobic bacteria and aerobic ammonia-oxidizing archaea (AOA). The oxygen-sensitive bacterial 4HBD and oxygen-tolerant archaeal 4HBD share 59 % amino acid identity. To examine the structural basis of oxygen tolerance in archaeal 4HBD, we determined the atomic resolution structure of the enzyme. Two tunnels providing access to the canonical [4Fe-4S] cluster in oxygen-sensitive bacterial 4HBD were closed with four conserved mutations found in all aerobic AOA and other archaea. Further biochemical experiments and molecular dynamics simulations support our findings that restricting access to the active site is the key to oxygen tolerance, explaining how active site evolution drove a major evolutionary transition. Significance statement Autotrophy (primary production) was the first life strategy on Earth. Before photosynthesis (using solar energy to fix carbon dioxide), life relied on chemical reactions for energy. These chemosynthetic reactions are present in all domains of life, including archaea possessing the most energy-efficient carbon fixation pathway - the 3-hydroxypropionate/4-hydroxybutyrate cycle. This efficiency results from enzyme modifications, including enhanced enzyme stability and catalysis of multiple reactions. We reveal the first structure of aerobic 4-hydroxybutyryl-CoA dehydratase (4HBD) from ammonia-oxidizing archaea. These archaea are among the most abundant organisms on the planet, and their 4HBD active site evolved oxygen tolerance to support aerobic metabolism. This modification can provide further insight into enzyme evolution on early earth, as photosynthesis developed and began oxygenating the atmosphere.
DeMirci, Hasan (ORCID:0000000291355397)↗