Identification of Dephospho-Coenzyme A (Dephospho-CoA) Kinase in <named-content content-type="genus-species">Thermococcus kodakarensis</named-content> and Elucidation of the Entire CoA Biosynthesis Pathway in Archaea

ABSTRACT Dephospho-coenzyme A (dephospho-CoA) kinase (DPCK) catalyzes the ATP-dependent phosphorylation of dephospho-CoA, the final step in coenzyme A (CoA) biosynthesis. DPCK has been identified and characterized in bacteria and eukaryotes but not in archaea. The hyperthermophilic archaeon Thermoco...

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Autores principales:	Takahiro Shimosaka, Kira S. Makarova, Eugene V. Koonin, Haruyuki Atomi
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Lenguaje:	EN
Publicado:	American Society for Microbiology 2019
Materias:	archaea coenzyme A dephospho-CoA kinase hyperthermophiles metabolism Microbiology QR1-502
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spelling	oai:doaj.org-article:b341fffb6f78494aabe0eb91a1534f952021-11-15T16:22:11ZIdentification of Dephospho-Coenzyme A (Dephospho-CoA) Kinase in <named-content content-type="genus-species">Thermococcus kodakarensis</named-content> and Elucidation of the Entire CoA Biosynthesis Pathway in Archaea10.1128/mBio.01146-192150-7511https://doaj.org/article/b341fffb6f78494aabe0eb91a1534f952019-08-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mBio.01146-19https://doaj.org/toc/2150-7511ABSTRACT Dephospho-coenzyme A (dephospho-CoA) kinase (DPCK) catalyzes the ATP-dependent phosphorylation of dephospho-CoA, the final step in coenzyme A (CoA) biosynthesis. DPCK has been identified and characterized in bacteria and eukaryotes but not in archaea. The hyperthermophilic archaeon Thermococcus kodakarensis encodes two homologs of bacterial DPCK and the DPCK domain of eukaryotic CoA synthase, TK1334 and TK2192. We purified the recombinant TK1334 and TK2192 proteins and found that they lacked DPCK activity. Bioinformatic analyses showed that, in several archaea, the uncharacterized gene from arCOG04076 protein is fused with the gene for phosphopantetheine adenylyltransferase (PPAT), which catalyzes the reaction upstream of the DPCK reaction in CoA biosynthesis. This observation suggested that members of arCOG04076, both fused to PPAT and standalone, could be the missing archaeal DPCKs. We purified the recombinant TK1697 protein, a standalone member of arCOG04076 from T. kodakarensis, and demonstrated its GTP-dependent DPCK activity. Disruption of the TK1697 resulted in CoA auxotrophy, indicating that TK1697 encodes a DPCK that contributes to CoA biosynthesis in T. kodakarensis. TK1697 homologs are widely distributed in archaea, suggesting that the arCOG04076 protein represents a novel family of DPCK that is not homologous to bacterial and eukaryotic DPCKs but is distantly related to bacterial and eukaryotic thiamine pyrophosphokinases. We also constructed and characterized gene disruption strains of TK0517 and TK2128, homologs of bifunctional phosphopantothenoylcysteine synthetase-phosphopantothenoylcysteine decarboxylase and PPAT, respectively. Both strains displayed CoA auxotrophy, indicating their contribution to CoA biosynthesis. Taken together with previous studies, the results experimentally validate the entire CoA biosynthesis pathway in T. kodakarensis. IMPORTANCE CoA is utilized in a wide range of metabolic pathways, and its biosynthesis is essential for all life. Pathways for CoA biosynthesis in bacteria and eukaryotes have been established. In archaea, however, the enzyme that catalyzes the final step in CoA biosynthesis, dephospho-CoA kinase (DPCK), had not been identified. In the present study, bioinformatic analyses identified a candidate for the DPCK in archaea, which was biochemically and genetically confirmed in the hyperthermophilic archaeon Thermococcus kodakarensis. Genetic analyses on genes presumed to encode bifunctional phosphopantothenoylcysteine synthetase-phosphopantothenoylcysteine decarboxylase and phosphopantetheine adenylyltransferase confirmed their involvement in CoA biosynthesis. Taken together with previous studies, the results reveal the entire pathway for CoA biosynthesis in a single archaeon and provide insight into the different mechanisms of CoA biosynthesis and their distribution in nature.Takahiro ShimosakaKira S. MakarovaEugene V. KooninHaruyuki AtomiAmerican Society for Microbiologyarticlearchaeacoenzyme Adephospho-CoA kinasehyperthermophilesmetabolismMicrobiologyQR1-502ENmBio, Vol 10, Iss 4 (2019)
institution	DOAJ
collection	DOAJ
language	EN
topic	archaea coenzyme A dephospho-CoA kinase hyperthermophiles metabolism Microbiology QR1-502
spellingShingle	archaea coenzyme A dephospho-CoA kinase hyperthermophiles metabolism Microbiology QR1-502 Takahiro Shimosaka Kira S. Makarova Eugene V. Koonin Haruyuki Atomi Identification of Dephospho-Coenzyme A (Dephospho-CoA) Kinase in <named-content content-type="genus-species">Thermococcus kodakarensis</named-content> and Elucidation of the Entire CoA Biosynthesis Pathway in Archaea
description	ABSTRACT Dephospho-coenzyme A (dephospho-CoA) kinase (DPCK) catalyzes the ATP-dependent phosphorylation of dephospho-CoA, the final step in coenzyme A (CoA) biosynthesis. DPCK has been identified and characterized in bacteria and eukaryotes but not in archaea. The hyperthermophilic archaeon Thermococcus kodakarensis encodes two homologs of bacterial DPCK and the DPCK domain of eukaryotic CoA synthase, TK1334 and TK2192. We purified the recombinant TK1334 and TK2192 proteins and found that they lacked DPCK activity. Bioinformatic analyses showed that, in several archaea, the uncharacterized gene from arCOG04076 protein is fused with the gene for phosphopantetheine adenylyltransferase (PPAT), which catalyzes the reaction upstream of the DPCK reaction in CoA biosynthesis. This observation suggested that members of arCOG04076, both fused to PPAT and standalone, could be the missing archaeal DPCKs. We purified the recombinant TK1697 protein, a standalone member of arCOG04076 from T. kodakarensis, and demonstrated its GTP-dependent DPCK activity. Disruption of the TK1697 resulted in CoA auxotrophy, indicating that TK1697 encodes a DPCK that contributes to CoA biosynthesis in T. kodakarensis. TK1697 homologs are widely distributed in archaea, suggesting that the arCOG04076 protein represents a novel family of DPCK that is not homologous to bacterial and eukaryotic DPCKs but is distantly related to bacterial and eukaryotic thiamine pyrophosphokinases. We also constructed and characterized gene disruption strains of TK0517 and TK2128, homologs of bifunctional phosphopantothenoylcysteine synthetase-phosphopantothenoylcysteine decarboxylase and PPAT, respectively. Both strains displayed CoA auxotrophy, indicating their contribution to CoA biosynthesis. Taken together with previous studies, the results experimentally validate the entire CoA biosynthesis pathway in T. kodakarensis. IMPORTANCE CoA is utilized in a wide range of metabolic pathways, and its biosynthesis is essential for all life. Pathways for CoA biosynthesis in bacteria and eukaryotes have been established. In archaea, however, the enzyme that catalyzes the final step in CoA biosynthesis, dephospho-CoA kinase (DPCK), had not been identified. In the present study, bioinformatic analyses identified a candidate for the DPCK in archaea, which was biochemically and genetically confirmed in the hyperthermophilic archaeon Thermococcus kodakarensis. Genetic analyses on genes presumed to encode bifunctional phosphopantothenoylcysteine synthetase-phosphopantothenoylcysteine decarboxylase and phosphopantetheine adenylyltransferase confirmed their involvement in CoA biosynthesis. Taken together with previous studies, the results reveal the entire pathway for CoA biosynthesis in a single archaeon and provide insight into the different mechanisms of CoA biosynthesis and their distribution in nature.
format	article
author	Takahiro Shimosaka Kira S. Makarova Eugene V. Koonin Haruyuki Atomi
author_facet	Takahiro Shimosaka Kira S. Makarova Eugene V. Koonin Haruyuki Atomi
author_sort	Takahiro Shimosaka
title	Identification of Dephospho-Coenzyme A (Dephospho-CoA) Kinase in <named-content content-type="genus-species">Thermococcus kodakarensis</named-content> and Elucidation of the Entire CoA Biosynthesis Pathway in Archaea
title_short	Identification of Dephospho-Coenzyme A (Dephospho-CoA) Kinase in <named-content content-type="genus-species">Thermococcus kodakarensis</named-content> and Elucidation of the Entire CoA Biosynthesis Pathway in Archaea
title_full	Identification of Dephospho-Coenzyme A (Dephospho-CoA) Kinase in <named-content content-type="genus-species">Thermococcus kodakarensis</named-content> and Elucidation of the Entire CoA Biosynthesis Pathway in Archaea
title_fullStr	Identification of Dephospho-Coenzyme A (Dephospho-CoA) Kinase in <named-content content-type="genus-species">Thermococcus kodakarensis</named-content> and Elucidation of the Entire CoA Biosynthesis Pathway in Archaea
title_full_unstemmed	Identification of Dephospho-Coenzyme A (Dephospho-CoA) Kinase in <named-content content-type="genus-species">Thermococcus kodakarensis</named-content> and Elucidation of the Entire CoA Biosynthesis Pathway in Archaea
title_sort	identification of dephospho-coenzyme a (dephospho-coa) kinase in <named-content content-type="genus-species">thermococcus kodakarensis</named-content> and elucidation of the entire coa biosynthesis pathway in archaea
publisher	American Society for Microbiology
publishDate	2019
url	https://doaj.org/article/b341fffb6f78494aabe0eb91a1534f95
work_keys_str_mv	AT takahiroshimosaka identificationofdephosphocoenzymeadephosphocoakinaseinnamedcontentcontenttypegenusspeciesthermococcuskodakarensisnamedcontentandelucidationoftheentirecoabiosynthesispathwayinarchaea AT kirasmakarova identificationofdephosphocoenzymeadephosphocoakinaseinnamedcontentcontenttypegenusspeciesthermococcuskodakarensisnamedcontentandelucidationoftheentirecoabiosynthesispathwayinarchaea AT eugenevkoonin identificationofdephosphocoenzymeadephosphocoakinaseinnamedcontentcontenttypegenusspeciesthermococcuskodakarensisnamedcontentandelucidationoftheentirecoabiosynthesispathwayinarchaea AT haruyukiatomi identificationofdephosphocoenzymeadephosphocoakinaseinnamedcontentcontenttypegenusspeciesthermococcuskodakarensisnamedcontentandelucidationoftheentirecoabiosynthesispathwayinarchaea
_version_	1718426925676888064

Identification of Dephospho-Coenzyme A (Dephospho-CoA) Kinase in <named-content content-type="genus-species">Thermococcus kodakarensis</named-content> and Elucidation of the Entire CoA Biosynthesis Pathway in Archaea

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