Evidence of positive selection on D-lactate dehydrogenases in Lactobacillus delbrueckii subsp. bulgaricus

Abstract

Lactobacillus delbrueckii has been widely used in combination with Streptococcus thermophilus for yogurt fermentation. It has genes encoding both D- and L-type lactate dehydrogenase (LDH) that catalyze the production of L (+) or D (−) stereoisomer of lactic acid, respectively. D-lactic acid is the primary lactate product by L. delbrueckii, yet it cannot be metabolized by human intestine. Since L. delbrueckii has been domesticated for long time, an interesting question arises regarding to whether the selection pressure has affected the evolution of both L-LDH and D-LDH genes in the genome. To answer this question, in this study we first investigated the evolution of these two genes by constructing phylogenetic trees from the orthologous LDH genes of 48 lactate producing species. We found that D-LDH-based phylogenetic tree could better represent the phylogenetic relationship between species including L. delbrueckii in the acidophilus complex than L-LDH-based tree, indicating that it could serve a molecular marker for phylogenetic studies of the acidophilus complex. We next investigated the evolution of both D-LDH and L-LDH genes at amino acid level, and found that D-LDH gene in L. delbrueckii is positively selected, possibly a consequence of long-term domestication. We further identified four amino acids in D-LDH gene that are under positive selection. One of the positively selected residues, V261, is located in the center of three catalytic active sites, and is approximate to the binding sites of D-LDH in three-dimensional structure, indicating likely functional effects on the enzyme activity. The selection from the domestication process thus provides direction for future engineering of D-LDH.