Purification of a Crenarchaeal ATP Synthase in the Light of the Unique Bioenergetics of Species

Overview

Journal J Bacteriol

Publisher American Society for Microbiology

Specialty Microbiology

Date 2019 Jan 16

PMID 30642991

Authors

Lydia J Kreuter

Andrea Weinfurtner

Alexander Ziegler

Julia Weigl

Jan Hoffmann

Nina Morgner

Volker Muller

Harald Huber

Affiliations

Soon will be listed here.

Abstract

In this study, the ATP synthase of was purified, characterized, and structurally compared to the respective enzymes of the other species, to shed light on energy conservation in this unique group of archaea. The crenarchaeal genus comprises three described species, i.e., and from hot marine sediments near Iceland and from a hydrothermal vent system in the Pacific Ocean. This genus is unique among all archaea due to the unusual cell envelope, consisting of two membranes that enclose a large intermembrane compartment (IMC). is the best studied member of this genus, mainly because it is the only known host for the potentially parasitic archaeon grows chemolithoautotrophically, and its sole energy-yielding reaction is the reduction of elemental sulfur with molecular hydrogen, forming large amounts of hydrogen sulfide. This reaction generates an electrochemical gradient, which is used by the ATP synthase, located in the outer cellular membrane, to generate ATP inside the IMC. The genome of encodes nine subunits of an A-type ATP synthase, which we could identify in the purified complex. Although the maximal activity of the enzyme was measured around pH 6, the optimal stability of the AA complex seemed to be at pH 9. Interestingly, the soluble A subcomplexes of the different species exhibited significant differences in their apparent molecular masses in native electrophoresis, although their behaviors in gel filtration and chromatography-mass spectrometry were very similar. The represent one of the major phyla within the domain. This study describes the successful purification of a crenarchaeal ATP synthase. To date, all information about A-type ATP synthases is from euryarchaeal enzymes. The fact that it has not been possible to purify this enzyme complex from a member of the until now points to significant differences in stability, possibly caused by structural alterations. Furthermore, the study subject has a particular importance among crenarchaeotes, since it is the only known host of The energy metabolism in this system is still poorly understood, and our results can help elucidate the unique relationship between these two microbes.

References

Stark H . GraFix: stabilization of fragile macromolecular complexes for single particle cryo-EM. Methods Enzymol. 2010; 481:109-26. DOI: 10.1016/S0076-6879(10)81005-5. View

Schagger H . Tricine-SDS-PAGE. Nat Protoc. 2007; 1(1):16-22. DOI: 10.1038/nprot.2006.4. View

Gruber G, Subramanian Manimekalai M, Mayer F, Muller V . ATP synthases from archaea: the beauty of a molecular motor. Biochim Biophys Acta. 2014; 1837(6):940-52. DOI: 10.1016/j.bbabio.2014.03.004. View

Jahn U, Gallenberger M, Paper W, Junglas B, Eisenreich W, Stetter K . Nanoarchaeum equitans and Ignicoccus hospitalis: new insights into a unique, intimate association of two archaea. J Bacteriol. 2008; 190(5):1743-50. PMC: 2258681. DOI: 10.1128/JB.01731-07. View

Nass B, Poll U, Langer J, Kreuter L, Kuper U, Flechsler J . Three multihaem cytochromes c from the hyperthermophilic archaeon Ignicoccus hospitalis: purification, properties and localization. Microbiology (Reading). 2014; 160(Pt 6):1278-1289. DOI: 10.1099/mic.0.077792-0. View

Hilario E, Gogarten J . Horizontal transfer of ATPase genes--the tree of life becomes a net of life. Biosystems. 1993; 31(2-3):111-9. DOI: 10.1016/0303-2647(93)90038-e. View

Cipriano D, Wang Y, Bond S, Hinton A, Jefferies K, Qi J . Structure and regulation of the vacuolar ATPases. Biochim Biophys Acta. 2008; 1777(7-8):599-604. PMC: 2467516. DOI: 10.1016/j.bbabio.2008.03.013. View

Sobott F, Hernandez H, McCammon M, Tito M, Robinson C . A tandem mass spectrometer for improved transmission and analysis of large macromolecular assemblies. Anal Chem. 2002; 74(6):1402-7. DOI: 10.1021/ac0110552. View

Rachel R, Wyschkony I, Riehl S, Huber H . The ultrastructure of Ignicoccus: evidence for a novel outer membrane and for intracellular vesicle budding in an archaeon. Archaea. 2005; 1(1):9-18. PMC: 2685547. DOI: 10.1155/2002/307480. View

10.

Muller V, Gruber G . ATP synthases: structure, function and evolution of unique energy converters. Cell Mol Life Sci. 2003; 60(3):474-94. PMC: 11138706. DOI: 10.1007/s000180300040. View

11.

Wilms R, Freiberg C, Wegerle E, Meier I, Mayer F, Muller V . Subunit structure and organization of the genes of the A1A0 ATPase from the Archaeon Methanosarcina mazei Gö1. J Biol Chem. 1996; 271(31):18843-52. DOI: 10.1074/jbc.271.31.18843. View

12.

Morgner N, Robinson C . Massign: an assignment strategy for maximizing information from the mass spectra of heterogeneous protein assemblies. Anal Chem. 2012; 84(6):2939-48. DOI: 10.1021/ac300056a. View

13.

Giannone R, Huber H, Karpinets T, Heimerl T, Kuper U, Rachel R . Proteomic characterization of cellular and molecular processes that enable the Nanoarchaeum equitans--Ignicoccus hospitalis relationship. PLoS One. 2011; 6(8):e22942. PMC: 3149612. DOI: 10.1371/journal.pone.0022942. View

14.

Suhai T, Heidrich N, Dencher N, Seelert H . Highly sensitive detection of ATPase activity in native gels. Electrophoresis. 2009; 30(20):3622-5. DOI: 10.1002/elps.200900114. View

15.

BOYER P . The ATP synthase--a splendid molecular machine. Annu Rev Biochem. 1997; 66:717-49. DOI: 10.1146/annurev.biochem.66.1.717. View

16.

Kastner B, Fischer N, Golas M, Sander B, Dube P, Boehringer D . GraFix: sample preparation for single-particle electron cryomicroscopy. Nat Methods. 2007; 5(1):53-5. DOI: 10.1038/nmeth1139. View

17.

Vonck J, Pisa K, Morgner N, Brutschy B, Muller V . Three-dimensional structure of A1A0 ATP synthase from the hyperthermophilic archaeon Pyrococcus furiosus by electron microscopy. J Biol Chem. 2009; 284(15):10110-9. PMC: 2665065. DOI: 10.1074/jbc.M808498200. View

18.

Kuper U, Meyer C, Muller V, Rachel R, Huber H . Energized outer membrane and spatial separation of metabolic processes in the hyperthermophilic Archaeon Ignicoccus hospitalis. Proc Natl Acad Sci U S A. 2010; 107(7):3152-6. PMC: 2840320. DOI: 10.1073/pnas.0911711107. View

19.

Lewalter K, Muller V . Bioenergetics of archaea: ancient energy conserving mechanisms developed in the early history of life. Biochim Biophys Acta. 2006; 1757(5-6):437-45. DOI: 10.1016/j.bbabio.2006.04.027. View

20.

Konishi J, Wakagi T, Oshima T, Yoshida M . Purification and properties of the ATPase solubilized from membranes of an acidothermophilic archaebacterium, Sulfolobus acidocaldarius. J Biochem. 1987; 102(6):1379-87. DOI: 10.1093/oxfordjournals.jbchem.a122184. View