8qmp

Publication Abstract from PubMed

The autoinhibited plasma membrane calcium ATPase ACA8 from A. thaliana has an N-terminal autoinhibitory domain. Binding of calcium-loaded calmodulin at two sites located at residues 42-62 and 74-96 relieves autoinhibition of ACA8 activity. Through activity studies and a yeast complementation assay we investigated wild-type (WT) and N-terminally truncated ACA8 constructs (Delta20, Delta30, Delta35, Delta37, Delta40, Delta74 and Delta100) to explore the role of conserved motifs in the N-terminal segment preceding the calmodulin binding sites. Furthermore, we purified WT, Delta20- and Delta100-ACA8, tested activity in vitro and performed structural studies of purified Delta20-ACA8 stabilized in a lipid nanodisc to explore the mechanism of autoinhibition. We show that an N-terminal segment between residues 20 and 35 including conserved Phe32, upstream of the calmodulin binding sites, is important for autoinhibition and the activation by calmodulin. Cryo-EM structure determination at 3.3 A resolution of a beryllium fluoride inhibited E2 form, and at low resolution for an E1 state combined with AlphaFold prediction provide a model for autoinhibition, consistent with the mutational studies.

Conserved N-terminal Regulation of the ACA8 Calcium Pump with Two Calmodulin Binding Sites.,Larsen ST, Dannerso JK, Nielsen CJF, Poulsen LR, Palmgren M, Nissen P J Mol Biol. 2024 Oct 15;436(20):168747. doi: 10.1016/j.jmb.2024.168747. Epub 2024 , Aug 20. PMID:39168442^[1]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.