TY - JOUR
T1 - Ca2+ induces spontaneous dephosphorylation of a novel P5A-type ATPase
AU - Sørensen, Danny Mollerup
AU - Møller, Annette B.
AU - Jakobsen, Mia K.
AU - Jensen, Michael Krogh
AU - Vangheluwe, Peter
AU - Buch-Pedersen, Morten J.
AU - Palmgren, Michael Broberg
PY - 2012/8/17
Y1 - 2012/8/17
N2 - P5 ATPases constitute the least studied group of P-type ATPases, an essential family of ion pumps in all kingdoms of life. Although P5 ATPases are present in every eukaryotic genome analyzed so far, they have remained orphan pumps, and their biochemical function is obscure. We show that a P5A ATPase from barley, HvP5A1, locates to the endoplasmic reticulum and is able to rescue knock-out mutants of P5A genes in both Arabidopsis thaliana and Saccharomyces cerevisiae. HvP5A1 spontaneously forms a phosphorylated reaction cycle intermediate at the catalytic residue Asp-488, whereas, among all plant nutrients tested, only Ca2+ triggers dephosphorylation. Remarkably, Ca2+-induced dephosphorylation occurs at high apparent [Ca 2+] (Ki = 0.25 mM) and is independent of the phosphatase motif of the pump and the putative binding site for transported ligands located in M4. Taken together, our results rule out that Ca2+ is a transported substrate but indicate the presence of a cytosolic low affinity Ca2+-binding site, which is conserved among P-type pumps and could be involved in pump regulation. Our work constitutes the first characterization of a P5 ATPase phosphoenzyme and points to Ca2+ as a modifier of its function.
AB - P5 ATPases constitute the least studied group of P-type ATPases, an essential family of ion pumps in all kingdoms of life. Although P5 ATPases are present in every eukaryotic genome analyzed so far, they have remained orphan pumps, and their biochemical function is obscure. We show that a P5A ATPase from barley, HvP5A1, locates to the endoplasmic reticulum and is able to rescue knock-out mutants of P5A genes in both Arabidopsis thaliana and Saccharomyces cerevisiae. HvP5A1 spontaneously forms a phosphorylated reaction cycle intermediate at the catalytic residue Asp-488, whereas, among all plant nutrients tested, only Ca2+ triggers dephosphorylation. Remarkably, Ca2+-induced dephosphorylation occurs at high apparent [Ca 2+] (Ki = 0.25 mM) and is independent of the phosphatase motif of the pump and the putative binding site for transported ligands located in M4. Taken together, our results rule out that Ca2+ is a transported substrate but indicate the presence of a cytosolic low affinity Ca2+-binding site, which is conserved among P-type pumps and could be involved in pump regulation. Our work constitutes the first characterization of a P5 ATPase phosphoenzyme and points to Ca2+ as a modifier of its function.
U2 - 10.1074/jbc.m112.387191
DO - 10.1074/jbc.m112.387191
M3 - Journal article
C2 - 22730321
SN - 0021-9258
VL - 287
SP - 28336
EP - 28348
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
IS - 34
ER -