PT  - JOURNAL ARTICLE
AU  - Alexandra Bergman
AU  - Leonie Wenning
AU  - Verena Siewers
AU  - Jens Nielsen
TI  - Investigation of putative regulatory acetylation sites in Fas2p of <em>Saccharomyces cerevisiae</em>
AID  - 10.1101/430918
DP  - 2018 Jan 01
TA  - bioRxiv
PG  - 430918
4099  - http://biorxiv.org/content/early/2018/09/29/430918.short
4100  - http://biorxiv.org/content/early/2018/09/29/430918.full
AB  - Yeast metabolism is highly regulated, in part via coordinated reprogramming of metabolism on a transcriptional level, for example in response to environmental changes. Furthermore, regulation occurs on the protein level via posttranslational modifications directly affecting enzymatic activity – a mode of regulation that has the benefit of being very fast in response to environmental changes. One group of posttranslational modification that has been suggested to have a high impact on regulation of metabolism are acetylations. Around 4000 distinct protein acetylation sites have been found in Saccharomyces cerevisiae, many of which are located in central metabolic enzymes. However, reports on the verification of regulatory roles of specific acetylation sites on these metabolic enzymes have yet to emerge. This study investigates putative regulatory acetylation sites on Fas2p, which in concert with Fas1p is responsible for cytosolic fatty acid (FA) biosynthesis in S. cerevisiae. Fas2p stands out as one of the most highly acetylated proteins in yeast and is located at a branchpoint of acetyl-CoA metabolism. The amino acids (AAs) glutamine (Q) and arginine (R) were introduced to mimic a constitutively acetylated or non-acetylatable state at three separate lysine sites (K) (K83, K173 and K1551) confirmed to be acetylated in two independent studies, either separately or simultaneously. The results suggest that the residue replacement system in the specific case interferes with the enzymatic function of the fatty acid synthase (FAS), as QQQ and RRR triple mutants both reduce the amount of secreted free fatty acids (FFAs) in a faa1Δ faa4Δ yeast deletion mutant. The K173Q substitution significantly decreased C16 FA species at the expense of C18 FAs, while no such change could be observed for the corresponding K173R modification.