skip to main content

DOE PAGESDOE PAGES

Title: Flux-Enabled Exploration of the Role of Sip1 in Galactose Yeast Metabolism

13C metabolic flux analysis ( 13C MFA) is an important systems biology technique that has been used to investigate microbial metabolism for decades. The heterotrimer Snf1 kinase complex plays a key role in the preference Saccharomyces cerevisiae exhibits for glucose over galactose, a phenomenon known as glucose repression or carbon catabolite repression. The SIP1 gene, encoding a part of this complex, has received little attention, presumably, because its knockout lacks a growth phenotype. We present a fluxomic investigation of the relative effects of the presence of galactose in classically glucose-repressing media and/or knockout of SIP1 using a multi-scale variant of 13C MFA known as 2-Scale 13C metabolic flux analysis (2S- 13C MFA). In this study, all strains have the galactose metabolism deactivated (gal1Δ background) so as to be able to separate the metabolic effects purely related to glucose repression from those arising from galactose metabolism. The resulting flux profiles reveal that the presence of galactose in classically glucose-repressing conditions, for a CEN.PK113-7D gal1Δ background, results in a substantial decrease in pentose phosphate pathway (PPP) flux and increased flow from cytosolic pyruvate and malate through the mitochondria toward cytosolic branched-chain amino acid biosynthesis. These fluxomic redistributions are accompanied by a highermore » maximum specific growth rate, both seemingly in violation of glucose repression. Deletion of SIP1 in the CEN.PK113-7D gal1Δ cells grown in mixed glucose/galactose medium results in a further increase. Knockout of this gene in cells grown in glucose-only medium results in no change in growth rate and a corresponding decrease in glucose and ethanol exchange fluxes and flux through pathways involved in aspartate/threonine biosynthesis. Glucose repression appears to be violated at a 1/10 ratio of galactose-to-glucose. Based on the scientific literature, we may have conducted our experiments near a critical sugar ratio that is known to allow galactose to enter the cell. Additionally, we report a number of fluxomic changes associated with these growth rate increases and unexpected flux profile redistributions resulting from deletion of SIP1 in glucose-only medium.« less
Authors:
 [1] ;  [2] ;  [2] ;  [2] ;  [2] ;  [2] ;  [3] ;  [4]
  1. Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States); Joint BioEnergy Institute, Emeryville, CA (United States); Univ. of California, Berkeley, CA (United States)
  2. Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States); Joint BioEnergy Institute, Emeryville, CA (United States)
  3. Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States); Joint BioEnergy Institute, Emeryville, CA (United States); DOE Agile Biofoundry, Emeryville, CA (United States); Basque Center for Applied Mathematics, Bilbao, Basque Country (Spain)
  4. Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States); Joint BioEnergy Institute, Emeryville, CA (United States); Univ. of California, Berkeley, CA (United States); Technical Univ. of Denmark, Horsholm (Denmark)
Publication Date:
Grant/Contract Number:
AC02-05CH11231
Type:
Accepted Manuscript
Journal Name:
Frontiers in Bioengineering and Biotechnology
Additional Journal Information:
Journal Volume: 5; Journal ID: ISSN 2296-4185
Publisher:
Frontiers Research Foundation
Research Org:
Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States)
Sponsoring Org:
USDOE Office of Science (SC), Biological and Environmental Research (BER) (SC-23)
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; 60 APPLIED LIFE SCIENCES
OSTI Identifier:
1379658