Growth and energetics of Leuconostoc oenos during cometabolism of glucose with citrate or fructose

Abstract

The metabolic and energetic characterization of the growth of Leuconostoc oenos on glucose-citrate or glucose-fructose mixtures enables the potential role of this bacterium in the wine-making process to be ascertained. Moreover, mixotrophic conditions remain a suitable means for improving biomass productivities of malolactic starter cultures. When the malolactic bacterium L. oenos was grown in batch cultures on complex medium at pH 5.0 with glucose-citrate or glucose-fructose mixtures, enhancement of both the specific growth rate and biomass production yields was observed. While growth was possible on fructose as the sole source of energy, citrate alone did not allow subsequent biomass production. The metabolic interactions between the catabolic pathways of the glucose cosubstrates and the heterofermentation of hexoses led to an increased acetate yield as a result of modified NADH oxidation. However, the calculated global coenzyme regeneration showed that the reducing equivalent balance was never equilibrated. The stimulatory efects of these glucose cosubstrates on growth resulted from increased ATP synthesis by substrate-level phosphorylation via acetate kinase. While the energetic efficiency remained close to 10 g of biomass produced per mol of ATP, the increase in the specific growth rate and biomass production yields was directly related to the rate and yield of ATP generation.Leuconostoc oenos has the ability to convert L-( -)-malate to L-(+)-lactate and carbon dioxide in a single step catalyzed by L-malate:NAD' carboxylyase. This so-called malolactic fermentation is an important reaction in the vinification process, since it reduces acidity, enhances organoleptic characteristics, and improves the microbiological stability of wines (3). For several years, wine makers have tended to use selected malolactic bacteria to ensure better control of malic acid bioconversion in enology. Thus, studies have been performed to examine the use of L. oenos strains as malolactic starter cultures, with the aims of both optimizing growth and understanding better the metabolism of this bacterium.Glucose remains the most frequently employed catabolic substrate providing the energy necessary for the growth of Leuconostoc species. Hexoses are metabolized via the heterofermentative phosphoketolase pathway or the 6-phosphogluconate pathway, leading to equimolar amounts of lactate, C02, and ethanol-acetate (6). On the other hand, it is well known that a number of lactic acid bacteria, including Leuconostoc species, are able to metabolize citrate. Since citrate catabolism is of importance for the production of flavor compounds, such as diacetyl and acetoin, numerous studies have dealt with the cometabolism of citrate and sugars. However, many of these studies are difficult to interpret because the relationship between the substrates used and the products formed was not quantified. Recently, Cogan (2) described the kinetics of substrate utilization and product formation from glucose and glucose-citrate mixtures during the growth of se...

Malolactic fermentation in wine - beyond deacidification

“…The co-fermentation of citrate and glucose in O. oeni is physiologically important for this bacterium. This co-metabolism of citrate-glucose has been shown to enhance the growth rate and biomass yield of this bacterium, which result from increased ATP synthesis both by substrate-level phosphorylation via acetate kinase and by a chemiosmotic mechanism (proton motive force) (Salou et al 1994;Ramos and Santos 1996). The growth stimulation of citrate-sugar co-fermentation by the same mechanisms has also been reported in citrate-fermenting dairy LAB (Cogan 1987;Schmitt and Diviès 1991;Ramos et al 1994;Bandell et al 1998).…”

“…Wine contains a range of monosaccharides (pentoses and hexoses) and disaccharides, with arabinose, glucose, fructose and trehalose being the major sugars (Liu and Davis 1994). The utilization of sugars by wine LAB as carbon and energy sources during MLF has been demonstrated in a number of studies and there exist species and strain differences in sugar utilization (Davis et al 1986a, b;Salou et al 1994;Liu et al 1995a). Glucose and trehalose are generally preferred over other sugars (Liu 1990;Liu et al 1995a).…”

“…Being a ketose, fructose can also act as an electron acceptor and is reduced to mannitol. Consequently, acetyl phosphate formed during hexose fermentation is converted to acetate instead of being reduced to ethanol, generating an additional ATP (Pilone et al 1991;Salou et al 1994). Besides fructose, heterofermentative LAB can use other substances as electron acceptors, such as oxygen and pyruvate, resulting in the production of acetate and additional ATP.…”

Genomic analysis of PSU-1 and its relevance to winemaking

“…Numerous researchers have examined citrate utilization by O. oeni [7,29]. Like other LAB, O. oeni does not use citrate as a sole carbon source but co-metabolizes citrate with glucose resulting in higher achievable biomass than growth on glucose alone [30]. Upon transport into the cell, citrate is converted via the actions of citrate lyase and oxaloacetate decarboxylase to pyruvate, which is then converted to a mixture of lactate, acetate, diacetyl, acetoin, and 2,3-butanediol using a pathway similar to that found in other citrate fermenting dairy lactic acid bacteria [24].…”

Organic acid metabolism under different glucose concentrations of Leuconostoc oenos from wine

“…is able to metabolize citrate. Recently, Salou et a/. (1994) studied the co-metabolism of glucose with citrate or fructose by Leimnostoi.…”