Human glucose-6-phosphate dehydrogenase: the crystal structure reveals a structural NADP+ molecule and provides insights into enzyme deficiency

Abstract

Background: Glucose-6-phosphate dehydrogenase (G6PD) catalyses the first committed step in the pentose phosphate pathway; the generation of NADPH by this enzyme is essential for protection against oxidative stress. The human enzyme is in a dimer↔tetramer equilibrium and its stability is dependent on NADP+ concentration. G6PD deficiency results from many different point mutations in the X-linked gene encoding G6PD and is the most common human enzymopathy. Severe deficiency causes chronic non-spherocytic haemolytic anaemia; the usual symptoms are neonatal jaundice, favism and haemolytic anaemia.Results: We have determined the first crystal structure of a human G6PD (the mutant Canton, Arg459→Leu) at 3 Å resolution. The tetramer is a dimer of dimers. Despite very similar dimer topology, there are two major differences from G6PD of Leuconostoc mesenteroides: a structural NADP+ molecule, close to the dimer interface but integral to the subunit, is visible in all subunits of the human enzyme; and an intrasubunit disulphide bond tethers the otherwise disordered N-terminal segment. The few dimer–dimer contacts making the tetramer are charge–charge interactions.Conclusions: The importance of NADP+ for stability is explained by the structural NADP+ site, which is not conserved in prokaryotes. The structure shows that point mutations causing severe deficiency predominate close to the structural NADP+ and the dimer interface, primarily affecting the stability of the molecule. They also indicate that a stable dimer is essential to retain activity in vivo. As there is an absolute requirement for some G6PD activity, residues essential for coenzyme or substrate binding are rarely modified.