Mechanism of surcrose utilisation by Saccharomyces cerevisiae.

Abstract

Saccharomyces cerevisiae was grown on different concentrations of sucrose and glucose mixtures after adapting on sucrose. The yeast cells were found to have two different mechanisms by which sucrose was utilised: hydrolysis outside the cell membrane and direct transport into the cells. The mechanism by which sucrose was utilised depended on the initial concentration of glucose in the mixture and the adaptation state of the cells. In both cases, glucose was utilised first and invertase secretion was repressed when the glucose concentration was higher than 2 g l-'. The major important finding in our results is that, for fully sucrose-adapted cells, even in the presence of a repressive glucose concentration, the yeast cells were able to utilise sucrose, and most importantly, without first being hydrolysed to its constituent monosaccharides. Previously, it has been thought that direct transport of sucrose contributes only a small part of the process of utilisation of sucrose, but as our results indicate it can be the major mechanism of sucrose utilisation in S. cerevisiae if the cells are adapted on sucrose for long periods, glucose concentrations in the medium are high and/or invertase activities are limiting.The initial steps in the utilisation of sugars by yeast is either the passage of intact sugar molecules across the cell membrane or the hydrolysis outside the cell followed by entry of all or some of the hydrolytic products into the cell. Sugar transport into microbial cells across the plasmalemma can take place by active transport, facilitated diffusion and group translocation (19).Sucrose utilisation involves its hydrolysis outside the yeast cell by extracellular enzyme invertase ($-D-fructosidase) into glucose and fructose which are then transported into the cells and metabolised. Most Saccharomyces cerevisiae strains have two forms of invertase: an external large invertase, which is a glycoprotein, and an internal small invertase (9,15). Some strains of S. cerevisiae have been reported to transport sucrose directly into the cell (14,17,20).

Active α-glucoside transport in Saccharomyces cerevisiae

“…While there is no doubt about the importance of the maltose and K-glucoside permeases for the brewing and baking industries, in the case of distilleries and fuel ethanol industries which are based on the fermentation of the sucrose-rich sugar cane, the Kglucoside symporter encoded by the AGT1 permease is of greater signi¢cance, as it is probably responsible for the previously characterized [12] sucrose-H symport activity of yeast cells. Indeed, recent studies have demonstrated that the observed fermentation and growth rates on sucrose can only ¢t a model in which both the direct uptake of sucrose and the uptake of its hydrolysis products contribute to the total uptake [13]. Although it is generally accepted that the ¢rst step in sucrose utilization is its hydrol-ysis by periplasmic invertase, a transport activity would also justify the existence of a constitutive intracellular invertase which arises from a shorter transcript of the same gene (e.g.…”

Active α-glucoside transport inSaccharomyces cerevisiae

“…While there is no doubt about the importance of the maltose and α‐glucoside permeases for the brewing and baking industries, in the case of distilleries and fuel ethanol industries which are based on the fermentation of the sucrose‐rich sugar cane, the α‐glucoside symporter encoded by the AGT1 permease is of greater significance, as it is probably responsible for the previously characterized [12] sucrose‐H + symport activity of yeast cells. Indeed, recent studies have demonstrated that the observed fermentation and growth rates on sucrose can only fit a model in which both the direct uptake of sucrose and the uptake of its hydrolysis products contribute to the total uptake [13]. Although it is generally accepted that the first step in sucrose utilization is its hydrolysis by periplasmic invertase, a transport activity would also justify the existence of a constitutive intracellular invertase which arises from a shorter transcript of the same gene (e.g.…”

Efeito da alteração na captação de sacarose ao metabolismo de Saccharomyces cerevisiae.

“…A característica dessa ligação confere a esse carboidrato, a possibilidade de sofrer a ação de dois tipos de hidrolases: por α-D-glicosidases, que são capazes de reconhecer a ligação "Gli α1", e por β-D-frutosidases, que são capazes de reconhecer a ligação "β2 Fru". Dessa forma, na levedura S. cerevisiae são conhecidas duas vias de utilização para esse açúcar, demonstrando haver inter-relação do metabolismo da sacarose com de outros carboidratos: a sacarose pode ser hidrolisada no ambiente extracelular -em glicose e frutose -por ação da invertase extracelular, e os monossacarídeos produzidos pela hidrólise da sacarose são captados por transportadores HXT; e também, conforme ilustra a Figura 2, a sacarose pode ser captada diretamente para o interior das células através do co-transporte com prótons H + , com alta afinidade pelo transportador codificado pelo gene AGT1 e com baixa afinidade pelos transportadores MALT, e posteriormente hidrolisada pela maltase (α-D-glicosidase) ou pela invertase (β-Dfrutosidase) intracelulares STAMBUK, 2006;BADOTTI et al, 2008;PHILLIPS;ORLOWSKI, 1992;STAMBUK, 2004;MWESIGYE;BARFORD, 1996;BARFORD, 1991 ZIMMERMANN, 1979;HOHMANN;ZIMMERMANN, 1986;KORSHUNOVA;NAUMOV, 2005;NAUMOV et al, 1996;NAUMOVA, 2010a,b). A análise de leveduras, isoladas em diferentes ambientes industriais, revelou que S. cerevisiae apresenta alta atividade invertase e também vários loci SUC no genoma, quando expostas a meios de cultura onde a sacarose é o principal açúcar a ser fermentado (como caldo-de-cana ou melaço).…”

“…Em cervejaria, os açúcares presentes no mosto (maltose e maltotriose) são captados ativamente pelas células e hidrolisados no citoplasma, fato que permite um equilíbrio entre as concentrações dos açúcares no interior e no exterior da célula, impedindo alteração drástica na osmolaridade do meio (KODAMA et al, 1994(KODAMA et al, , 1995ZASTROW et al, 2001). Trabalhos demonstram que a sacarose é também metabolizada por transporte direto através da membrana, para o interior das células, e hidrólise pela invertase intracelular (BADOTTI et al, 2008;PHILLIPS;ORLOWSKI, 1992;DÁRIO, 2007;MWESIGYE;BARFORD, 1996). Por outro lado, a metabolização desse dissacarídeo ocorre principalmente por hidrólise extracelular, por ação da invertase extracelular (DÁRIO, 2007;CARLSON, 1992).…”