Structural basis of cofactor-mediated stabilization and substrate recognition of the α-tubulin acetyltransferase αTAT1

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Abstract

The functions of microtubules are controlled in part by tubulin post-translational modification including acetylation of Lys40 in α-tubulin. αTAT1 (α-tubulin acetyltransferase 1), an enzyme evolutionarily conserved among eukaryotes, has recently been identified as the major α-tubulin Lys40 acetyltransferase, in which AcCoA (acetyl-CoA) serves as an acetyl group donor. The regulation and substrate recognition of this enzyme, however, have not been fully understood. In the present study, we show that AcCoA and CoA each form a stable complex with human αTAT1 to maintain the protein integrity both in vivo and in vitro. The invariant residues Arg132 and Ser160 in αTAT1 participate in the stable interaction not only with AcCoA but also with CoA, which is supported by analysis of the present crystal structures of the αTAT1 catalytic domain in complex with CoA. Alanine substitution for Arg132 or Ser160 leads to a drastic misfolding of the isolated αTAT1 catalytic domain in the absence of CoA and AcCoA but not in the presence of excess amounts of either cofactor. A mutant αTAT1 carrying the R132A or S160A substitution is degraded much faster than the wild-type protein when expressed in mammalian Madin-Darby canine kidney cells. Furthermore, alanine-scanning experiments using Lys40-containing peptides reveal that α-tubulin Ser38 is crucial for substrate recognition of αTAT1, whereas Asp39, Ile42, the glycine stretch (amino acid residues 43-45) and Asp46 are also involved. The requirement for substrate selection is totally different from that in various histone acetyltransferases, which appears to be consistent with the inability of αTAT1 to acetylate histones.

Original languageEnglish
Pages (from-to)103-113
Number of pages11
JournalBiochemical Journal
Volume467
Issue number1
DOIs
Publication statusPublished - Apr 1 2015

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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