The optimal temperature for growth of L. lactis subsp. Chemicals. It is worth noting that our main goal was not the optimization of mannitol … Interestingly, the relative proportions of the reduced products derived from pyruvate (lactate, ethanol, and 2,3-butanediol) were similar in both strains, but the acetoin/2,3-butanediol ratio was considerably enhanced in the mannitol-producing mutant. Furthermore, insight into how the transcription of the mannitol operon in L. lactis is regulated will provide useful clues for manipulating mannitol production in this organism. After its inactivation, 60% of the glucose was converted into mannitol by cells suspended in glucose buffer. <>/Font<>/ProcSet[/PDF/Text/ImageB]/XObject<>>>/Rotate 0/TrimBox[9 9 594 792]/Type/Page>> uuid:f709f47f-1dd1-11b2-0a00-b80000000000
Specific PCR fragments used for construction of deletion vectors were amplified by using Bio-X-Act DNA polymerase (Bioline).

lactis strains from various environments. Ligation reactions were performed according to Sambrook et al.
We also observed that the mannitol produced was taken up and rapidly metabolized after glucose depletion; therefore, it was apparent that the design of a mannitol-producing strain would have to consider the ability of L. lactis to utilize mannitol as an energy source for growth (27). <>/Font<>/ProcSet[/PDF/Text]>>/Rotate 0/TrimBox[9 9 594 792]/Type/Page>> The ATP demand for anabolic processes could limit mannitol synthesis during growth. Restriction enzymes and other DNA-modifying enzymes were used according to the suppliers' recommendations. 1) (3); today it is classified as Lactococcus lactis subsp. In all of the experiments performed with nongrowing cells, the carbon and redox balances were on average 95% ± 2% and 98% ± 3%, respectively. Assuming a similar Km for L. lactis Mtl1PDH, the F6P concentration could constitute a limiting factor for mannitol production, since the F6P pools are generally low. Users are For the quantitative analysis of the NMR sample extracts by 13C-NMR, a repetition delay of 60.5 s was used. We noted unexpected but significant differences in the growth behaviors of both strains. Construction of gene replacement vectors. Moreover, the yield of mannitol can be further improved by implementing genetic manipulations along the lines discussed above. , using a slightly different engineering strategy. Growth was evaluated by measuring the turbidity (optical density of the culture at 600 nm [OD600]) and calibrating against cell dry mass measurements. The double mutants, in contrast to the parent strain, were unable to utilize mannitol even after glucose depletion, showing that mannitol was taken up exclusively by PEP-PTSMtl. Copyright © 2020 American Society for Microbiology | Privacy Policy | Website feedback, Print ISSN: 0099-2240; Online ISSN: 1098-5336, Sign In to Email Alerts with your Email Address. 4B). DMPy: a Python package for automated mathematical model construction of large-scale metabolic systems. This article is cited by It has been yet reported that in some cases, lactis strains derived from different sources was as follows: marine fish‐derived strain > cheese starter‐derived strain > freshwater fish‐derived strain (Fig. endobj Growth on glucose yielded formate, ethanol, and acetate as the main end products. Comparisons of the product yields derived from the metabolism of glucose during growth and in nongrowing cells are presented in Table 3. Most of the lactococci resemble the enterococci in presumptive tests (BE, NaCl, PYR, LAP). lactis strains from different sources.