Pedersen, Bjørn P. , Nissen, Poul , Buch-Pedersen, Morten J. , Palmgren, Michael G. .
An X-ray crystal structure of a plant plasma membrane H+-ATPase.
The plasma membrane (PM) H+-ATPase is the major active transport system in fungi1 and plants2,3 where it extrudes protons from cells and in this way energizes the PM. The transmembrane electrochemical gradient of protons in turn provides the driving force for nutrient uptake through channel proteins and secondary active transport systems. By forming a phosphorylated reaction cycle intermediate4 the plant PM H+-ATPase displays the hallmark of P-type ATPases, a large ubiquitous family of cation pumps5. So far, the only pump of this family for which structures are present is the rabbit sarco/endoplasmatic reticulum Ca2+-ATPase (SERCA1)6,7.
Using the yeast Saccharomyces cerevisiae as a heterologous expression system we produced large amounts of recombinant Arabidopsis thaliana autoinhibited PM H+-ATPase isoform 2 (AHA2). In the presence of a nucleotide analogue to inhibit pump dynamics, and by employing a mutated pump (D684N) devoid of flexible terminal domains and frozen in the E1 conformation, we were able to obtain a crystal structure of the pump using data extending to 3.5 Ã… resolution. The crystal structures show a single polypeptide protein of ten transmembrane helices and three cytosolic domains (N, P and A). Residues provide liganding oxygens for intramembraneous binding of protons in the transmembrane region, a regulatory K+ in the cytoplasmic region and a divalent cation at the apoplastic side of the pump. The cytosolic nucleotide binding (N) domain likely serves as a protein kinase that start each catalytic cycle by phosphorylating the phosphorylation (P) domain, which in turn is dephosphorylated by the phosphatase (A) domain. Large domain motions are hypothesized to be coupled to twisting and pulling of membrane helices, which in turn generate entrance and exit pathways for protons.
The structure of the plant PM H+-ATPase is remarkable similar to that of the Ca2+-ATPase, even in transmembrane regions where no strong sequence similarity exists between the two pumps. Further, the structure is in support of the previous findings8 that Asp-684 in transmembrane segment six is an essential residue for proton coordination. This strongly suggests that protons are transported by essentially the same mechanism as Ca2+ by the Ca2+-ATPase.
1Serrano R, Kielland-Brandt MC, Fink GR (1986) Nature 319:689-693.
2Harper JF, Surowy TK, Sussman MR (1989) Proc Natl Acad Sci U S A 86:1234-1238.
3Pardo JM, Serrano R (1989) J Biol Chem 264:8557-62.
4Vara F, Serrano R (1983) J Biol Chem 258:5334-5336.
5Axelsen KB, Palmgren MG (1998) J Mol Evol 46:84-101.
6Toyoshima C, Nakasako M, Nomura H, Ogawa H (2000) Nature 405:647-655.
7Olesen C, Sorensen TL, Nielsen RC, Moller JV, Nissen P (2004) Science 306:2251-2255.
8Buch-Pedersen MJ, Palmgren MG (2003) J Biol Chem 278:17845-17851.
Log in to add this item to your schedule
1 - University of Aarhus, Denmark, Dept Structural Biology
2 - University of Copenhagen, Denmark, Dept Plant Biology
3 - University of Copenhagen, Denmark, Dept Plant Biology, 40, Thorvaldsensvej, Frederiksberg C, DK-1871, Frederiksberg C
Presentation Type: Plant Biology Abstract
Location: Exhibit Hall (Northeast, Southwest & Southeast)/Hilton
Date: Sunday, July 8th, 2007
Time: 8:00 AM