First evidence of changes in enzyme kinetics and stability of glucokinase affected by somatic cancer-associated variations

• Published in: Biochimica et biophysica acta. Proteins and proteomics Vol. 1867; no. 3; pp. 213 - 218
• Main Authors: Těšínský, Miroslav, Šimčíková, Daniela, Heneberg, Petr
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 01.03.2019
• Subjects: active sites; carcinogenesis; Connecting region I; energy metabolism; enzyme kinetics; genome; glucokinase; glucose; glycolysis; half life; Hexokinase D; Lung cancer; Melanoma; N-acetylglucosamine; neoplasm cells; neoplasms; Persistent hypoglycemic hyperinsulinemia of infancy; phosphorylation; recombinant proteins; Somatic nonsynonymous substitutions; thermal stability; G6P; GSIR-T; PNDM; Hexokinase D; BAD; Lung cancer; Melanoma; GST; Connecting region I; GCK; MODY; COSMIC; Persistent hypoglycemic hyperinsulinemia of infancy; GlcNAc; RAI; PHHI; Somatic nonsynonymous substitutions
• ISSN: 1570-9639; 1878-1454; 1878-1454
• DOI: 10.1016/j.bbapap.2018.12.008

Recent investigation of somatic variations of allosterically regulated proteins in cancer genomes suggested that variations in glucokinase (GCK) might play a role in tumorigenesis. We hypothesized that somatic cancer-associated GCK variations include in part those with activating and/or stabilizing effects. We analyzed the enzyme kinetics and thermostability of recombinant proteins possessing the likely activating variations and the variations present in the connecting loop I and provided the first experimental evidence of the effects of somatic cancer-associated GCK variations. Activating and/or stabilizing variations were common among the analyzed cancer-associated variations, which was in strong contrast to their low frequency among germinal variations. The activating and stabilizing variations displayed focal distribution with respect to the tertiary structure, and were present in the surroundings of the heterotropic allosteric activator site, including but not limited to the connecting loop I and in the active site region subject to extensive rearrangements upon glucose binding. Activating somatic cancer-associated variations induced a reduction of GCK's cooperativity and an increase in the affinity to glucose (a decline in the S0.5 values). The hotspot-associated variations, which decreased cooperativity, also increased the half-maximal inhibitory concentrations of the competitive GCK inhibitor, N-acetylglucosamine. Concluded, we have provided the first convincing biochemical evidence establishing GCK as a previously unrecognized enzyme that contributes to the reprogramming of energy metabolism in cancer cells. Activating GCK variations substantially increase affinity of GCK to glucose, disrupt the otherwise characteristic sigmoidal response to glucose and/or prolong the enzyme half-life. This, combined, facilitates glucose phosphorylation, thus supporting glycolysis and associated pathways. [Display omitted] •Somatic GCK variations are known particularly from skin and colorectal cancers•GCK contributes to the reprogramming of energy metabolism in cancer cells•Activating somatic GCK variations overlap with PHHI-inducing germinal variations•Ability for non-cooperative binding of glucose is often disrupted•IC50 of the competitive GCK inhibitor N-acetylglucosamine is often increased