Analytical and Bioanalytical Chemistry

, Volume 376, Issue 7, pp 994–1005 | Cite as

Acetylation of the HIV-1 Tat protein: an in vitro study

  • Wilma Dormeyer
  • Alexander Dorr
  • Melanie Ott
  • Martina Schnölzer
Special Issue Paper

Abstract

In the last few years, the understanding of lysine acetylation as a regulatory post-translational modification of proteins in cell signalling cascades has increased. It is now known that not only histones but also non-histone factors can serve as substrates of different acetyltransferase enzymes. Acetylated lysine residues in non-histone factors are often identified using radioactive labelling experiments and immunochemical analysis of synthetic peptides. In this study of the human immunodeficiency virus 1 (HIV-1) Tat protein, we demonstrate the benefits of matrix-assisted laser desorption/ionisation mass spectrometry, proteolytic digestion and Edman sequencing for the mapping of acetylation sites. We confirmed that the HIV-1 Tat protein is acetylated in vitro by the acetyltransferase p300 at a specific lysine residue at position 50 in its RNA binding region. Furthermore, we showed that the Tat cysteine-rich region is acetylated at multiple cysteine residues in the absence of enzyme. Since this non-enzymatic cysteine acetylation occurs independently from the surrounding peptide sequence, we consider the presence of cysteine residues in acetylated peptides an important factor for the interpretation of in vitro acetylation assays in general.

Keywords

Acetylation MALDI-TOF mass spectrometry Edman sequencing HIV-1 Tat Histone acetyltransferase p300 

Abbreviations

aa

Amino acid

AcCoA

Acetyl coenzyme A

acm

Acetamidomethyl

ARM

Arginine-rich motif

CRR

Cysteine-rich region

HAT

Histone acetyltransferase

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Copyright information

© Springer-Verlag 2003

Authors and Affiliations

  • Wilma Dormeyer
    • 1
  • Alexander Dorr
    • 2
  • Melanie Ott
    • 2
  • Martina Schnölzer
    • 1
  1. 1.Protein Analysis FacilityDeutsches Krebsforschungszentrum (DKFZ)HeidelbergGermany
  2. 2.Applied Tumour VirologyDeutsches Krebsforschungszentrum (DKFZ) HeidelbergGermany

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