Disruption of the sole ldhL gene in Lactobacillus sakei prevents the production of both L- and D-lactate.

Abstract

A 7 kb DNA fragment was cloned from Lactobacillus sakei which contains the IdhL gene encoding the L(+)-lactate dehydrogenase (L-LDH). Analysis of the DNA sequence, Northern experiments and primer extension experiments showed that IdhL is transcribed from a single promoter, leading to a monocistronic 1.15 kb mRNA which yields the L-LDH. A stable mutant was constructed by chromosomal integration of a chloramphenicol cassette into IdhL by a double-crossover event. Both L- and D-lactate were produced by the wild-type strain whereas only residual amounts of both isomers were produced by the mutant. This demonstrates that L. sakei possesses an L-LDH producing L-lactate and a lactate racemase able to transform it to D-lactate, but is devoid of D-LDH activity. Moreover the ability to degrade L-lactate present in the medium that was observed with the mutant strain grown aerobically suggests that an L-lactate oxidase activity is also present in L. sakei.