AUTHOR=VanDrisse Chelsey M. , Escalante-Semerena Jorge C. 

TITLE=In Salmonella enterica, OatA (Formerly YjgM) Uses O-Acetyl-Serine and Acetyl-CoA to Synthesize N,O-Diacetylserine, Which Upregulates Cysteine Biosynthesis

JOURNAL=Frontiers in Microbiology

VOLUME=9

YEAR=2018

URL=https://www.frontiersin.org/journals/microbiology/articles/10.3389/fmicb.2018.02838

DOI=10.3389/fmicb.2018.02838

ISSN=1664-302X

ABSTRACT=<p><sc>L</sc>-Cysteine biosynthesis has been extensively analyzed in <italic>Salmonella enterica</italic>. The cysteine regulon contains the genes whose protein products are necessary to convert sulfate to sulfide, which is eventually reacted with <italic>O</italic>-acetyl-serine (OAS) to generate cysteine. The LysR type regulator, CysB, is required for activation of the cysteine regulon, and its interaction with various <italic>cys</italic> genes has been thoroughly characterized. Results from previous studies by others, suggested that OAS undergoes a spontaneous <italic>O</italic>- to <italic>N-</italic> migration to produce <italic>N</italic>-acetyl-serine (NAS), and that NAS is the true signal sensed by CysB. It was unclear, however, whether such migration occurred spontaneously <italic>in vivo</italic> or if NAS was generated enzymatically. Work reported herein characterizes a <italic>S. enterica N</italic>-acetyltransferase, OatA (formerly YjgM), which acetylates the <italic>N</italic><sub>α</sub>-amino group of OAS, producing <italic>N,O-</italic>diacetyl-serine (DAS) at the expense of acetyl-CoA. We isolated OatA to homogeneity and performed its initial biochemical characterization. The product of the OatA reaction was isolated by HPLC and confirmed by mass spectrometry to be DAS; OatA did not acetylate NAS, consistent with the conclusion that OatA is an <italic>N-</italic>acetyltransferase, not an <italic>O-</italic>acetyltransferase. Binding of OAS to OatA appears to be positively cooperative with the apparent <italic>K<sub>0.5</sub></italic> for OAS determined to be 0.74 mM, the <italic>k<sub>cat</sub></italic> was 1.05 s<sup>-1</sup>, and the catalytic efficiency of the enzyme (<italic>k</italic><sub>cat</sub>/<italic>K<sub>0.5</sub></italic>) was 1.4 × 10<sup>3</sup> M<sup>-1</sup> s<sup>-1</sup>. Size exclusion chromatography indicated that OatA was a monomer in solution. In <italic>S. enterica</italic>, overexpression of <italic>oatA</italic> led to shorter lag times on sulfate-limiting medium and that these delayed lag times were due to increased expression of the cysteine regulon, as indicated by RT-qPCR results. OatA is the first Gcn5-related <italic>N-</italic>acetyltransferase (aka GNAT) involved in the regulation of amino acid biosynthetic genes in <italic>Salmonella</italic>. On the basis of results of transcriptomics studies performed by other investigators, we hypothesize that DAS may play a role in biofilm formation in <italic>S. enterica</italic> and other bacteria.</p>