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Isoforms of the DHTKD1-Encoded 2-Oxoadipate Dehydrogenase, Identified in Animal Tissues, Are not Observed upon the Human DHTKD1 Expression in Bacterial or Yeast Systems

A. I. Boyko1,a*, A. V. Artiukhov1,2, T. Kaehne3, M. L. di Salvo4, M. C. Bonaccorsi di Patti4, R. Contestabile4, A. Tramonti4,5, and V. I. Bunik1,2,6,b*

1Faculty of Bioengineering and Bioinformatics, Lomonosov Moscow State University, 119234 Moscow, Russia

2Belozersky Institute of Physico-Chemical Biology, Lomonosov Moscow State University, 119234 Moscow, Russia

3Institute of Experimental Internal Medicine, Otto-von-Guericke University, 39120 Magdeburg, Germany

4Department of Biological Sciences A. Rossi Fanelli, Sapienza University, 00185 Rome, Italy

5Institute of Molecular Biology and Pathology, Council of National Research, 00185 Rome, Italy

6Department of Biological Chemistry, Sechenov First Moscow State Medical University, 119146 Moscow, Russia

* To whom correspondence should be addressed.

Received June 16, 2020; Revised June 25, 2020; Accepted June 26, 2020
Unlike the OGDH-encoded 2-oxoglutarate dehydrogenase (OGDH), which is an essential enzyme present in all animal tissues, expression of the DHTKD1-encoded isoenzyme, 2-oxoadipate dehydrogenase (OADH), depends on a number of factors, and mutant DHTKD1 phenotypes are rarely manifested. Physiological significance of OADH is also obscured by the fact that both isoenzymes transform 2-oxoglutarate and 2-oxoadipate. By analogy with other members of the 2-oxo acid dehydrogenases family, OADH is assumed to be a component of the multienzyme complex that catalyzes oxidative decarboxylation of 2-oxoadipate. This study aims at molecular characterization of OADH from animal tissues. Phylogenetic analysis of 2-oxo acid dehydrogenases reveals OADH only in animals and Dictyostelium discoideum slime mold, within a common branch with bacterial OGDH. Examination of partially purified animal OADH by immunoblotting and mass spectrometry identifies two OADH isoforms with molecular weights of about 130 and 70 kDa. These isoforms are not observed upon the expression of human DHTKD1 protein in either bacterial or yeast system, where the synthesized OADH is of expected molecular weight (about 100 kDa). Thus, the OADH isoforms present in animal tissues, may result from the animal-specific regulation of the DHTKD1 expression and/or posttranslational modifications of the encoded protein. Mapping of the peptides identified in the OADH preparations, onto the protein structure suggests that the 70-kDa isoform is truncated at the N-terminus, but retains the active site. Since the N-terminal domain of OGDH is required for the formation of the multienzyme complex, it is possible that the 70-kDa isoform catalyzes non-oxidative transformation of dicarboxylic 2-oxo acids that does not require the multienzyme structure. In this case, the ratio of the OADH isoforms in animal tissues may correspond to the ratio between the oxidative and non-oxidative decarboxylation of 2-oxoadipate.
KEY WORDS: DHTKD1, OGDH, carboligase, 2-oxoglutarate dehydrogenase isoenzyme, 2-oxoadipate dehydrogenase isoform, posttranslational modifications

DOI: 10.1134/S0006297920080076