Insights into human phosphoglycerate kinase 1 deficiency as a conformational disease from biochemical, biophysical, and in vitro expression analyses

Abstract

Mutations in genes encoding metabolic enzymes are often the cause of inherited diseases. Mutations usually affect the ability of proteins to fold properly, thus leading to enzyme loss of function. In this work, we explored the relationships between protein stability, aggregation, and degradation in vitro and inside cells in a large set of mutants associated with human phosphoglycerate kinase 1 (hPGK1) deficiency. To this end, we studied a third of the pathogenic alleles reported in the literature using expression analyses and biochemical, biophysical, and computational procedures. Our results show that most pathogenic variants studied had an increased tendency to aggregate when expressed in Escherichia coli, well correlating with the denaturation half-lives measured by thermal denaturation in vitro. Further, the most deleterious mutants show reduced stability toward chemical denaturation and proteolysis, supporting a pivotal role of thermodynamic stability in the propensity toward aggregation and proteolysis of pathogenic hPGK1 mutants in vitro and inside cells. Our strategy allowed us to unravel the complex relationships between protein stability, aggregation, and degradation in hPGK1 deficiency, which might be used to understand disease mechanisms in many inborn errors of metabolism. Our results suggest that pharmacological chaperones and protein homeostasis modulators could be considered as good candidates for therapeutic approaches for hPGK1 deficiency.

Site-to-site interdomain communication may mediate different loss-of-function mechanisms in a cancer-associated NQO1 polymorphism

“…Due to the high prevalence of missense mutations affecting protein stability, activity and regulation1921464748, our mutational strategy combined with functional, structural, energetic and dynamic studies could help to decipher complex disease-associated mutational effects in many other loss-of-function genetic diseases and to identify structural targets for therapeutic correction.…”

Phosphoglycerate kinase deficiency: A nationwide multicenter retrospective study

“…However, these findings were observed in a small series of patients, and more experiments are needed to corroborate the correlation between phenotype and biochemical defect. From a molecular viewpoint, several findings indicate that the clinical manifestations associated with PGK deficiency may depend on the distinctive type of perturbations caused by mutations in the PGK1 gene . For instance, it has been suggested that mutations critically impairing protein stability preferentially provoke hemolytic anemia and neurological disorders while mutations impairing both catalytic efficiency and heat stability preferentially provoke isolated myopathy .…”

“…For instance, it has been suggested that mutations critically impairing protein stability preferentially provoke hemolytic anemia and neurological disorders while mutations impairing both catalytic efficiency and heat stability preferentially provoke isolated myopathy . Also, it has been shown that most PGK1 deleterious mutations display reduced stability toward chemical denaturation and proteolysis, suggesting a crucial role of thermodynamic stability in the propensity of PGK mutants to aggregate in vitro and in vivo . However, as several exceptions to these correlations have been described, the clinical symptoms cannot be understood only on the bases of molecular properties of the mutant enzyme .…”

Metabolic enzymes of helminth parasites: potential as drug targets

“…These diseases tend to be rare, but the symptoms are often severe. Examples include triose phosphate isomerase deficiency (OMIM #615512), phosphoglycerate kinase 1 deficiency (OMIM # 300653), hereditary fructose intolerance (aldolase B deficiency; OMIM #22960) and classic galactosemia (galactose 1-phosphate uridylyltransferase deficiency; OMIM #230400) [27][28][29][30][31]. In addition to reduced energy production, these diseases often have additional phenotypes.…”