The aim of this study was to evaluate the competency of Lactiplantibacillus plantarum 12-3 isolated from Tibetan kefir grains that how it converts linoleic acid (LA) into fatty acid metabolites and what are the main reactions involved in it. Also, we scrutinize the enzymes involved in this study via density functional theory (DFT) and in silico approaches. The taxonomic identity was performed using average nucleotide identity (ANI) analysis and to investigate its genome properties using the rapid annotations using subsystems technology (RAST) annotation service. After eliminating plasmid sequences to focus on core genomic information, ANI analysis was performed using the JSpecies Web Server. The results verified L. plantarum 12-3’s categorization as a member of the L. plantarum species, demonstrating good conservation and taxonomic relatedness. Heatmapper was used to visualize the ANI data clustering and heatmap, allowing the discovery of closely related strains within L. plantarum. RAST annotation of the genome revealed functional subsystems as well as metabolic pathways, cellular activities, and virulence factors. Several routes of future research might be pursued to further investigate the possible applications and distinctive properties of the L. plantarum 12-3 strain. To begin, comparative genomics studies with other L. plantarum strains would provide a better knowledge of the strain’s distinctive genetic variants and evolutionary adaptations. This may give light on its applicability for a variety of industrial uses, including food fermentation and probiotics.