3pdt

Publication Abstract from PubMed

Dictyostelium discoideum myosin II heavy chain kinase A (MHCK A), a member of the atypical alpha-kinase family, phosphorylates sites in the myosin II tail that block filament assembly. Here we show that the catalytic activity of A-CAT, the alpha-kinase domain of MHCK A (residues 552-841), is severely inhibited by the removal of a disordered C-terminal tail sequence (C-tail; residues 806-841). The key residue in the C-tail was identified as Thr825, which was found to be constitutively autophosphorylated. Dephosphorylation of Thr825 using shrimp alkaline phosphatase decreased A-CAT activity. The activity of a truncated ACAT lacking Thr825 could be rescued by Pi, phosphothreonine and a phosphorylated peptide, but not by threonine, glutamic acid, aspartic acid or an unphosphorylated peptide. These results focused attention on a Pi-binding pocket located in the C-terminal lobe of A-CAT. Mutational analysis demonstrated that the Pi-pocket was essential for A-CAT activity. Based on these results, it is proposed that autophosphorylation of Thr825 activates ACAT by providing a covalently-tethered ligand for the Pi-pocket. Ab initio modeling studies using the Rosetta FloppyTail and FlexPepDock protocols showed that it is feasible for the phosphorylated Thr825 to dock intramolecularly into the Pi-pocket. Allosteric activation is predicted to involve a conformational change in Arg734, which bridges the bound Pi to Asp762 in a key active site loop. Sequence alignments indicate that a comparable regulatory mechanism is likely to be conserved in Dictyostelium MHCK B-D and metazoan eukaryotic elongation factor-2 kinases.

Autophosphorylation activates Dictyostelium myosin II heavy chain kinase A by providing a ligand for an allosteric binding site in the {alpha}-kinase domain.,Crawley SW, Samimi Gharaei M, Ye Q, Yang Y, Raveh B, London N, Schueler-Furman O, Jia Z, Cote GP J Biol Chem. 2010 Nov 11. PMID:21071445^[1]

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