Exploring the genetic information of Lactobacillus helveticus strains hydrolyzing soy protein using genome-wide association analysis

Soy protein is a multimeric globular protein that is less likely to be metabolized and utilized by lactic acid bacteria (LAB) during fermentation compared to milk-derived proteins. Little genetic information is available regarding LAB-mediated hydrolysis of soy protein. In this study, we sequenced the genomes of 46  Lactobacillus helveticus ( L. helveticus ) strains to analyze genetic-level factors affecting soy protein hydrolytic activity. The analysis revealed that the soy protein hydrolysis capability of these L. helveticus strains showed no correlation with their isolation sources, geographic origins, extracellular protease genes, peptide transporter genes, or peptidase gene distribution. Through genome-wide association analysis (GWAS), we identified two significantly associated missense SNPs (SNP-482062 and SNP-1780770) linked to soy protein hydrolysis activity. These two SNPs changed the local tertiary structure of amino acid permease and ornithine decarboxylase, respectively. Enzymatic assays confirmed that the mutations significantly affected enzyme activity, thereby validating their critical role in soy protein hydrolysis. This study elucidates the genetic basis for differential soy protein hydrolysis capacity among L. helveticus strains, offering practical guidance for selecting highly proteolytic strains to develop soy-based fermented dairy alternatives.