TY - JOUR
T1 - Characterization and X-ray structure of the NADH-dependent coenzyme A disulfide reductase from Thermus thermophilus
AU - Lencina, Andrea M.
AU - Koepke, Juergen
AU - Preu, Julia
AU - Muenke, Cornelia
AU - Gennis, Robert B.
AU - Michel, Hartmut
AU - Schurig-Briccio, Lici A.
N1 - Funding Information:
We thank all members of the Gennis and Michel laboratories for help and useful discussions. We thank Dr. Claiborne (Wake Forest School of Medicine) for kindly providing the E. coli strain and plasmid expressing the CoADR from B. anthracis and for insightful scientific discussions and expertise. We also thank to Dr. Yagi (Scripps Research Institute, La Jolla) for providing us experimental information. We thank to Dr. Parsonage (Wake Forest School of Medicine) and Dr. Crane (Pomona College) for providing the E. coli strains and plasmids expressing the CoADRs from P. horikoshii and S. aureus , respectively. We thank Dr. G.N. Bennett (Rice University, Houston) for the GNB10608 strain. This work was supported by grants from the United States National Institutes of Health GM095600 and HL16101 to R.B.G. and from the Max Planck Society and the Deutsche Forschungsgemeinschaft (Cluster of Excellence “Macromolecular Complexes” Frankfurt) to H.M.
Funding Information:
We thank all members of the Gennis and Michel laboratories for help and useful discussions. We thank Dr. Claiborne (Wake Forest School of Medicine) for kindly providing the E. coli strain and plasmid expressing the CoADR from B. anthracis and for insightful scientific discussions and expertise. We also thank to Dr. Yagi (Scripps Research Institute, La Jolla) for providing us experimental information. We thank to Dr. Parsonage (Wake Forest School of Medicine) and Dr. Crane (Pomona College) for providing the E. coli strains and plasmids expressing the CoADRs from P. horikoshii and S. aureus, respectively. We thank Dr. G.N. Bennett (Rice University, Houston) for the GNB10608 strain. This work was supported by grants from the United States National Institutes of Health GM095600 and HL16101 to R.B.G. and from the Max Planck Society and the Deutsche Forschungsgemeinschaft (Cluster of Excellence ?Macromolecular Complexes? Frankfurt) to H.M.
Publisher Copyright:
© 2019 Elsevier B.V.
PY - 2019/11/1
Y1 - 2019/11/1
N2 - The crystal structure of the enzyme previously characterized as a type-2 NADH:menaquinone oxidoreductase (NDH-2) from Thermus thermophilus has been solved at a resolution of 2.9 Å and revealed that this protein is, in fact, a coenzyme A-disulfide reductase (CoADR). Coenzyme A (CoASH) replaces glutathione as the major low molecular weight thiol in Thermus thermophilus and is maintained in the reduced state by this enzyme (CoADR). Although the enzyme does exhibit NADH:menadione oxidoreductase activity expected for NDH-2 enzymes, the specific activity with CoAD as an electron acceptor is about 5-fold higher than with menadione. Furthermore, the crystal structure contains coenzyme A covalently linked Cys44, a catalytic intermediate (Cys44-S-S-CoA) reduced by NADH via the FAD cofactor. Soaking the crystals with menadione shows that menadione can bind to a site near the redox active FAD, consistent with the observed NADH:menadione oxidoreductase activity. CoADRs from other species were also examined and shown to have measurable NADH:menadione oxidoreductase activity. Although a common feature of this family of enzymes, no biological relevance is proposed. The CoADR from T. thermophilus is a soluble homodimeric enzyme. Expression of the recombinant TtCoADR at high levels in E. coli results in a small fraction that co-purifies with the membrane fraction, which was used previously to isolate the enzyme wrongly identified as a membrane-bound NDH-2. It is concluded that T. thermophilus does not contain an authentic NDH-2 component in its aerobic respiratory chain.
AB - The crystal structure of the enzyme previously characterized as a type-2 NADH:menaquinone oxidoreductase (NDH-2) from Thermus thermophilus has been solved at a resolution of 2.9 Å and revealed that this protein is, in fact, a coenzyme A-disulfide reductase (CoADR). Coenzyme A (CoASH) replaces glutathione as the major low molecular weight thiol in Thermus thermophilus and is maintained in the reduced state by this enzyme (CoADR). Although the enzyme does exhibit NADH:menadione oxidoreductase activity expected for NDH-2 enzymes, the specific activity with CoAD as an electron acceptor is about 5-fold higher than with menadione. Furthermore, the crystal structure contains coenzyme A covalently linked Cys44, a catalytic intermediate (Cys44-S-S-CoA) reduced by NADH via the FAD cofactor. Soaking the crystals with menadione shows that menadione can bind to a site near the redox active FAD, consistent with the observed NADH:menadione oxidoreductase activity. CoADRs from other species were also examined and shown to have measurable NADH:menadione oxidoreductase activity. Although a common feature of this family of enzymes, no biological relevance is proposed. The CoADR from T. thermophilus is a soluble homodimeric enzyme. Expression of the recombinant TtCoADR at high levels in E. coli results in a small fraction that co-purifies with the membrane fraction, which was used previously to isolate the enzyme wrongly identified as a membrane-bound NDH-2. It is concluded that T. thermophilus does not contain an authentic NDH-2 component in its aerobic respiratory chain.
KW - Coenzyme A disulfide reductase
KW - Flavoprotein
KW - NADH oxidation
KW - Thermus thermophilus
KW - X-ray structure
UR - http://www.scopus.com/inward/record.url?scp=85073121218&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=85073121218&partnerID=8YFLogxK
U2 - 10.1016/j.bbabio.2019.148080
DO - 10.1016/j.bbabio.2019.148080
M3 - Article
C2 - 31520616
AN - SCOPUS:85073121218
VL - 1860
JO - Biochimica et Biophysica Acta - Bioenergetics
JF - Biochimica et Biophysica Acta - Bioenergetics
SN - 0005-2728
IS - 11
M1 - 148080
ER -