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Properties of the pyridoxaldimine form of glutamate semialdehyde aminotransferase (glutamate-1-semialdehyde 2,1-aminomutase) and analysis of its role as an intermediate in the formation of aminolaevulinate.

ABSTRACT: Glutamate semialdehyde aminotransferase (glutamate-1-semialdehyde 2,1-aminomutase; EC 5.4.3.8) was converted into its pyridoxaldimine form by exhaustive replacement of endogenous pyridoxamine phosphate with pyridoxal phosphate. The isomerization of glutamate 1-semialdehyde to 5-aminolaevulinate by this form of the enzyme followed an accelerating time course which indicated that the enzyme initially had no activity but was converted into the active pyridoxamine phosphate form in an exponential process characterized by a rate constant (k) of 0.027 s-1. The pyridoxaldimine form of the enzyme was converted rapidly into the pyridoxamine form by (S)-4-aminohex-5-enoate and much more slowly by 4-aminobutyrate. The steady-state velocity of the enzyme increased in a markedly non-linear fashion with increasing enzyme concentration, indicating that the extent of dissociation of an intermediate in the reaction to free diaminovalerate and the pyridoxaldimine form of the enzyme depends upon the concentration of the enzyme.

SUBMITTER: Tyacke RJ

PROVIDER: S-EPMC1134422 | biostudies-other | 1993 Aug

REPOSITORIES: biostudies-other

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Project description:BACKGROUND:Bioenergetic characterisation of malignant tissues revealed that different tumour cells can catabolise multiple substrates as salvage pathways, in response to metabolic stress. Altered metabolism in gliomas has received a lot of attention, especially in relation to IDH mutations, and the associated oncometabolite D-2-hydroxyglutarate (2-HG) that impact on metabolism, epigenetics and redox status. Astrocytomas and oligodendrogliomas, collectively called diffuse gliomas, are derived from astrocytes and oligodendrocytes that are in metabolic symbiosis with neurons; astrocytes can catabolise neuron-derived glutamate and gamma-aminobutyric acid (GABA) for supporting and regulating neuronal functions. METHODS:Metabolic characteristics of human glioma cell models - including mitochondrial function, glycolytic pathway and energy substrate oxidation - in relation to IDH mutation status and after 2-HG incubation were studied to understand the Janus-faced role of IDH1 mutations in the progression of gliomas/astrocytomas. The metabolic and bioenergetic features were identified in glioma cells using wild-type and genetically engineered IDH1-mutant glioblastoma cell lines by metabolic analyses with Seahorse, protein expression studies and liquid chromatography-mass spectrometry. RESULTS:U251 glioma cells were characterised by high levels of glutamine, glutamate and GABA oxidation. Succinic semialdehyde dehydrogenase (SSADH) expression was correlated to GABA oxidation. GABA addition to glioma cells increased proliferation rates. Expression of mutated IDH1 and treatment with 2-HG reduced glutamine and GABA oxidation, diminished the pro-proliferative effect of GABA in SSADH expressing cells. SSADH protein overexpression was found in almost all studied human cases with no significant association between SSADH expression and clinicopathological parameters (e.g. IDH mutation). CONCLUSIONS:Our findings demonstrate that SSADH expression may participate in the oxidation and/or consumption of GABA in gliomas, furthermore, GABA oxidation capacity may contribute to proliferation and worse prognosis of gliomas. Moreover, IDH mutation and 2-HG production inhibit GABA oxidation in glioma cells. Based on these data, GABA oxidation and SSADH activity could be additional therapeutic targets in gliomas/glioblastomas.

Dataset Information

Properties of the pyridoxaldimine form of glutamate semialdehyde aminotransferase (glutamate-1-semialdehyde 2,1-aminomutase) and analysis of its role as an intermediate in the formation of aminolaevulinate.

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