N-terminal cysteine acetylation and oxidation patterns may define protein stability

Area of Science:

• Biochemistry
• Molecular Biology
• Cellular Physiology

Background:

• Oxygen homeostasis is crucial, maintained by O2-sensing enzymes that trigger adaptive responses to hypoxia.
• The enzyme ADO targets proteins like RGS4/5 and IL32 for degradation via the Cysteine/Arginine N-degron pathway by oxidizing N-terminal cysteines.
• The identification of additional ADO substrates is limited, potentially due to competing N-terminal cysteine modifications like acetylation.

Purpose of the Study:

• To investigate the interplay between ADO-catalyzed oxidation and NatA-catalyzed acetylation of N-terminal cysteine residues.
• To identify sequence determinants that direct either oxidation or acetylation of N-terminal cysteines.
• To understand the implications for oxygen-dependent protein stability and the hypoxic response.

Main Methods:

• In vitro and in vivo experiments to assess N-terminal cysteine acetylation by human NatA.
• Analysis of protein sequences to determine substrate specificity for ADO (oxidation) and NatA (acetylation).
• Investigating the mutual exclusivity of these two modifications.

Main Results:

• Human NatA catalyzes N-terminal cysteine acetylation both in vitro and in vivo.
• Downstream amino acid sequences dictate whether N-terminal cysteines are oxidized by ADO (preferring basic/aromatic residues) or acetylated by NatA (preferring acidic/polar residues).
• The modifications appear mutually exclusive, suggesting distinct protein pools.

Conclusions:

• Sequence context is a key determinant for N-terminal cysteine modification by oxidation or acetylation.
• These findings shed light on the regulation of protein stability in response to oxygen levels.
• The study provides insights into the mechanisms governing the hypoxic response.