The use of polylactic acid (PLA) as a biodegradable carbohydrate-based plas­tic is rapidly expanding in many areas such as food packaging, drug deliv­ery, textiles, medical implants, and cosmetics [111-114]. Both the physical properties and the rate of biodegradation can be controlled by adjusting the ratio of the blended enantiomers, D(-)-lactate and L(+)-lactate [115]. For decades, lactic acid bacteria have been used to produce optically pure d(-)- and L(+)-lactate. However, high costs due to the need for complex media and the inability to ferment a broad range of sugars have con­strained the use of PLA to the manufacture of medical grade sutures and implants. Alternative biocatalysts from a variety of organisms are currently being investigated for efficient and inexpensive production of optically pure isomers [116-121].

Biocatalysts derived from E. coli K-12 had been previously engineered to produce D(-)-lactate but these were not able to metabolize 10% glucose or sucrose to completion in rich or minimal media [122-124]. The success and robustness of E. coli W derived-ethanologenic KO11 prompted the redirection of metabolism in this organism from ethanol to D(-)-lactate production [39]. Elimination of adhE, ackA, and the Z. mobilis homoethanol pathway from KO11 yielded strain SZ110 [39]. SZ110 was subjected to metabolic evolution in LB 100 g L-1 glucose [39], mineral salts medium with 100 g L-1 sucrose, and mineral salts medium with 100 gL-1 glucose, along with genetic manipula­tions to reduce co-product formation and remove foreign genes, to ultimately generate D(-)-lactate-producer SZ194 [125]. Replacement of the native SZ194

IdhA gene with the Pediococcus acidilactici IdhL gene and further metabolic evolution in mineral salts medium with glucose resulted in L(+)-lactate pro­ducer strain TG103 [121]. Both SZ194 and TG103 produced 1.2 M lactate from 12% glucose in mineral salts medium supplemented with 1 mM betaine. How­ever, lactate optical purity decreased from 99.5% to 95% in the presence of betaine [42,121]. This chiral impurity was associated with high glycolytic flux rates; spillover of carbon to lactic acid through the methylglyoxal pathway was the source of the contamination [126-128]. Elimination of the first com­mitted enzyme of the methylglyoxal pathway (mgsA) restored the product optimal purity to close to 100% [121]. The resulting E. coli strains, TG114 and TG108, consistently produced high titers of greater than 99.9% chirally pure d(-)- and L(+)-lactate, respectively, from 12% glucose at greater than 95% of the theoretical yield [121]. The lactate titer, yield, and optical purity attained by TG114 and TG108 are the highest compared to other lactate-producing organisms and were achieved in simpler fermentation medium and condi­tion, making these microbial biocatalysts some of the most efficient lactate producers.

5.2