The dhod gene and deduced structure of mitochondrial dihydroorotate dehydrogenase in Engrailed melanogaster

Rawls, John M.; Kirkpatrick, Russell; Yang, Jun; Lacy, Laura R.

doi:10.1016/0378-1119(93)90393-h

Cited by 19 publications

(10 citation statements)

References 26 publications

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“…Clones containing cDNAs were prepared from previously described cDNAs for the DHODH proteins of Drosophila melanogaster [18], rat [19] and human [20]. Blunt ends of DNA molecules were created by treating restricted DNA with Klenow fragment of DNA polymerase I [21].…”

Section: Methodsmentioning

confidence: 99%

“…The pGEM4DM and pGEM4Mh clones contain the D. melanogaster [18] and human [20] DHODH ORFs, respectively, fused at their C‐terminal ends to the same enterokinase‐8×HIS tract ORF (derived from pASKCF) found in pGEM4C. Clones encoding the precursor polypeptides of the α‐subunit of matrix processing peptidase (α‐MPP), cytochrome c 1 heme lyase (CC 1 HL), and the fusion protein comprising the presequence of subunit 9 of F o ‐ATPase and dihydrofolate reductase (pSu9‐DHFR) were described earlier [24,25].…”

Section: Methodsmentioning

confidence: 99%

“…Escherichia coli ), but the nature of this association is unknown and this protein also lacks obvious localization sequences [15,16]. The N‐terminal portions of DHODH proteins found in plants, animals and other fungi contain motifs that resemble presequences and stop‐transfer sequences [14,17–19]. Experiments reported here analyze the roles of these N‐terminal sequences using yeast mitochondria as a model system to study the import requirements.…”

mentioning

confidence: 99%

“… N‐terminal sequences of DHODH proteins. The N‐termini of DHODH proteins of E. coli [16], Saccharomyces cerevisiae [14], Schizosaccharomyces pombe [14]; Arabidopsis thaliana [17]; Drosophila melanogaster [18], rat [19] and humans [20] are aligned, based upon a hydrophobic region (underlined) found in most eukaryotic proteins. An asterisk (*) indicates that the human sequence is incomplete at its extreme N‐terminus; however, the sequence shown has been demonstrated to direct mitochondrial localization of the protein [2].…”

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confidence: 99%

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Requirements for the mitochondrial import and localization of dihydroorotate dehydrogenase

Rawls

Knecht

Diekert

et al. 2000

European Journal of Biochemistry

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110

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In animals, dihydroorotate dehydrogenase (DHODH) is a mitochondrial protein that carries out the fourth step in de novo pyrimidine biosynthesis. Because this is the only enzyme of this pathway that is localized to mitochondria and because the enzyme is cytosolic in some bacteria and fungi, we carried out studies to understand the mode of targeting of animal DHODH and its submitochondrial localization. Analysis of fractionated rat liver mitochondria revealed that DHODH is an integral membrane protein exposed to the intermembrane space. In vitro-synthesized Drosophila, rat and human DHODH proteins were efficiently imported into the intermembrane space of isolated yeast mitochondria. Import did not alter the size of the in vitro synthesized protein, nor was there a detectable size difference when compared to the DHODH protein found in vivo. Thus, there is no apparent proteolytic processing of the protein during import either in vitro or in vivo. Import of rat DHODH into isolated yeast mitochondria required inner membrane potential and was at least partially dependent upon matrix ATP, indicating that its localization uses the well described import machinery of the mitochondrial inner membrane. The DHODH proteins of animals differ from the cytosolic proteins found in some bacteria and fungi by the presence of an N-terminal segment that resembles mitochondrial-targeting presequences. Deletion of the cationic portion of this N-terminal sequence from the rat DHODH protein blocked its import into isolated yeast mitochondria, whereas deletion of the adjacent hydrophobic segment resulted in import of the protein into the matrix. Thus, the N-terminus of the DHODH protein contains a bipartite signal that governs import and correct insertion into the mitochondrial inner membrane.

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

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confidence: 99%

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Requirements for the mitochondrial import and localization of dihydroorotate dehydrogenase

Rawls

Knecht

Diekert

et al. 2000

European Journal of Biochemistry

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110

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“…This is in contrast with many genes which are up-regulated or which have variants in the testis. For example, the angiotensin-converting-enzyme gene [42], the carbonic anhydrase II gene [43] and the Drosophila dihydro-orotate dehydrogenase gene [44] all use alternative testis-specific promoters.…”

Section: Figure 5 Effect Of Deletions In the Nrd-c Promoter Upon Lucimentioning

confidence: 99%

Gene expression of the dibasic-pair cleaving enzyme NRD convertase (N-arginine dibasic convertase) is differentially regulated in the GH3 pituitary and Mat-Lu prostate cell lines

Winter

Pierotti

2000

Biochem. J.

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Purification and characterization of dihydroorotate dehydrogenase a from lactococcus lactis, crystallization and preliminary X‐ray diffraction studies of the enzyme

et al. 1996

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Lactococcus lactis is the only organism known to contain two dihydroorotate dehydrogenases, i.e., the A-and B-forms. In this paper, we report the overproduction, purification, and crystallization of dihydroorotate dehydrogenase A. In solution, the enzyme is bright yellow. It is a dimer of subunits (34 kDa) that contain one molecule of flavin mononucleotide each. The enzyme shows optimal function in the pH range 7.5-9.0. It is specific for L-dihydroorotate as substrate and can use dichlorophenolindophenol, potassium hexacyanoferrate(III), and, to a lower extent, also molecular oxygen as acceptors of the reducing equivalents, whereas the pyridine nucleotide coenzymes (NAD+, NADP+) and the respiratory quinones (i.e., vitamins Q 6 , Qlo and K,) were inactive. The enzyme has been crystallized from solutions of 30% polyethylene glycol, 0.2 M sodium acetate, and 0.1 M Tris-HCI, pH 8.

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The dhod gene and deduced structure of mitochondrial dihydroorotate dehydrogenase in Engrailed melanogaster

Cited by 19 publications

References 26 publications

Requirements for the mitochondrial import and localization of dihydroorotate dehydrogenase

Requirements for the mitochondrial import and localization of dihydroorotate dehydrogenase

Gene expression of the dibasic-pair cleaving enzyme NRD convertase (N-arginine dibasic convertase) is differentially regulated in the GH3 pituitary and Mat-Lu prostate cell lines

Purification and characterization of dihydroorotate dehydrogenase a from lactococcus lactis, crystallization and preliminary X‐ray diffraction studies of the enzyme

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