The aminotransferase Aat initiates 3-phenyllactic acid biosynthesis in Pediococcus acidilactici

The function of the aminotransferase Aat (GenBank Protein WP_159211138) from Pediococcus acidilactici FAM 18098 was studied in vivo. For this purpose, the gene was replaced with an erythromycin resistance gene using the temperature-sensitive Escherichia coli-Pediococcus shuttle plasmid pSET4T_Delta aat. The knockout was verified by PCR and genome sequencing. Subsequently, the differences between the metabolism of the knockout and of the wild-type strain were investigated by determining the free amino acids and organic acids in culture supernatants. It was found that the knockout mutant no longer synthesized 3-phenyllactic acid (PLA) and 4-hydroxyphenyllactic acid (HPLA). Additionally, the mutant strain no longer catabolized phenylalanine. Metabolic pathway analysis using the KEGG database indicate that P. acidilactici cannot synthesize alpha-ketoglutarate that is a predominant amino-group acceptor in many transamination reactions. To study the transfer of the amino group of phenylalanine, the wild-type strain was incubated with [N-15] phenylalanine. Mass spectrometry showed that during fermentation, [N-15] alanine was formed, indicating that pyruvic acid is an amino group acceptor in P. acidilactici. The present study shows that Aat plays a crucial role in PLA/HPLA biosynthesis and pyruvic acid is an amino acceptor in transamination reactions in P. acidilactici.

Automated summary

The function of the Aat gene was studied in Pediococcus acidilactici FAM 18098 through gene knockout experiments. It was found that Aat plays a crucial role in the synthesis of 3-phenyllactic acid and 4-hydroxyphenyllactic acid, as well as the catabolism of phenylalanine. Pyruvic acid was identified as an amino group acceptor in transamination reactions in P. acidilactici. This study reveals the important metabolic function of Aat in P. acidilactici.