Cellular studies of the two main isoforms of human d-aspartate oxidase.
Valentina RabattoniLoredano PollegioniGabriella TedeschiElisa MaffioliSilvia SacchiPublished in: The FEBS journal (2021)
Human d-aspartate oxidase (hDASPO) is a FAD-dependent enzyme responsible for the degradation of d-aspartate (d-Asp). In the mammalian central nervous system, d-Asp behaves as a classical neurotransmitter, it is thought to be involved in neural development, brain morphology and behavior, and appears to be involved in several pathological states, such as schizophrenia and Alzheimer's disease. Apparently, the human DDO gene produces alternative transcripts encoding for three putative hDASPO isoforms, constituted by 341 (the 'canonical' form), 369, and 282 amino acids. Despite the increasing interest in hDASPO and its physiological role, little is known about these different isoforms. Here, the additional N-terminal peptide present in the hDASPO_369 isoform only has been identified in hippocampus of Alzheimer's disease female patients, while peptides corresponding to the remaining part of the protein were present in samples from male and female healthy controls and Alzheimer's disease patients. The hDASPO_369 isoform was largely expressed in E. coli as insoluble protein, hampering with its biochemical characterization. Furthermore, we generated U87 human glioblastoma cell clones stably expressing hDASPO_341 and, for the first time, hDASPO_369 isoforms; the latter protein showed a lower expression compared with the canonical isoform. Both protein isoforms are active (showing similar kinetic properties), localize to the peroxisomes, are very stable (a half-life of approximately 100 h has been estimated), and are primarily degraded through the ubiquitin-proteasome system. These studies shed light on the properties of hDASPO isoforms with the final aim to clarify the mechanisms controlling brain levels of the neuromodulator d-Asp.
Keyphrases
- endothelial cells
- amino acid
- end stage renal disease
- induced pluripotent stem cells
- ejection fraction
- newly diagnosed
- pluripotent stem cells
- chronic kidney disease
- binding protein
- cognitive decline
- protein protein
- stem cells
- peritoneal dialysis
- small molecule
- bipolar disorder
- multiple sclerosis
- cerebral ischemia
- dna methylation
- mesenchymal stem cells
- cerebrospinal fluid
- subarachnoid hemorrhage
- cognitive impairment