Paul Magnano:Week 3 Individual Journal

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*RNA analyses were done to see if RNA levels of nitrogen regulating genes changed with increased ammonia concentrations
*RNA analyses were done to see if RNA levels of nitrogen regulating genes changed with increased ammonia concentrations
*genes GDH1, GDH2,GLN1 were determined to study responses to ammonia levels
*genes GDH1, GDH2,GLN1 were determined to study responses to ammonia levels
 +
*GAP1 and PUT4 are amino acid permease genes
 +
*biosynthetic genes were ILV5 and HIS4
*Fig 2
*Fig 2
**increasing ammonia levels resulted in GDH1 levels staying the same
**increasing ammonia levels resulted in GDH1 levels staying the same
**ammonia concentrations of 29-44mM there was no GDH2, but with increase to 61mM GDH2 level increased
**ammonia concentrations of 29-44mM there was no GDH2, but with increase to 61mM GDH2 level increased
**maximum GLN1 expression was observed at 61mM
**maximum GLN1 expression was observed at 61mM
-
** overall it was seen that ammonia concentration both repressed GDH1 and caused the expression of GDH2
+
**overall it was seen that ammonia concentration both repressed GDH1 and caused the expression of GDH2
 +
**GAP1 was shown to be regulated in response to ammonia concentration
 +
**the influence of ammonia concentration on GAP1 and PUT4 was analzyed in this study
 +
**when ammonia concentration was above 44mM, GAP1 and PUT4 decreased
 +
**in ammonia limited culture a relationship between ammonia flux GAP1 expression wa observed
 +
**GAP1 and PUT4 expression are regulated by ammonia flux not by ammonia concentration
 +
**genes ILV5 and HIS4 increased with increasing ammonia concentration outside the cell, it was highest at 66mM
 +
 
 +
===Enzyme Activities===
==Ten New Terms==
==Ten New Terms==

Revision as of 23:19, 30 January 2013

Contents

Outline

Experiment

  • sacchromyces cerevisiae was grown in continuous culture, with varying ammonia input concentrations, while ammonia assiilation rates remained constant
  • ammonia is a preffered nitrogen source for s. cerevisiae, because it results in faster growth than other sources
  • in this study, cultures were grown with same flux (relationship b/w external ammonia concentration and rte of assimilation) but different inflow ammonia concentrations
  • Physiological parameters:
    • S. cerevisiae SU32 grown in continuous cultures with ammonia inflow concentrations ranging from 29 to 118mM
    • fixed glucose concentration of 100mM
  • Fig 1A
    • increase of ammonia concentration from 29 to 61mM resulted in increase of biomass from 4.9 to 8.2 g/liter
    • ammonia concentration above 61mM biomass remained at 8.2 g/liter
    • over trhe entire range of ammonia concentrations the ammonia flux into biomass was 1.1 mmol/gh
  • Fig 1B
    • input ammonia concentration above 44mM resulted in (c02 production/02 consumption) remained constant
    • with ammonia limitation (ammonia input below 44mM) c02 production and 02 consumption values differed
    • no changes in the residual glucose concentration were seen
    • no significant changes in carbon metabolism occured when the culture was switched from ammonia excess to ammonia limitation
  • Fig 1C
    • ammonia within the cells reacts with ketoglutarate to produce glutamate which is converted to glutamine
    • ketoglutarate concentration decreased from 10 to 5 umol/g when the ammonia concentration changed from limited to excess
    • glutamate concentration increased within the cell, linearly
    • increaing ammonia concentrations caused the concentrations of glutamate and glutamine within the cell to increase

Northern Analyses

  • RNA analyses were done to see if RNA levels of nitrogen regulating genes changed with increased ammonia concentrations
  • genes GDH1, GDH2,GLN1 were determined to study responses to ammonia levels
  • GAP1 and PUT4 are amino acid permease genes
  • biosynthetic genes were ILV5 and HIS4
  • Fig 2
    • increasing ammonia levels resulted in GDH1 levels staying the same
    • ammonia concentrations of 29-44mM there was no GDH2, but with increase to 61mM GDH2 level increased
    • maximum GLN1 expression was observed at 61mM
    • overall it was seen that ammonia concentration both repressed GDH1 and caused the expression of GDH2
    • GAP1 was shown to be regulated in response to ammonia concentration
    • the influence of ammonia concentration on GAP1 and PUT4 was analzyed in this study
    • when ammonia concentration was above 44mM, GAP1 and PUT4 decreased
    • in ammonia limited culture a relationship between ammonia flux GAP1 expression wa observed
    • GAP1 and PUT4 expression are regulated by ammonia flux not by ammonia concentration
    • genes ILV5 and HIS4 increased with increasing ammonia concentration outside the cell, it was highest at 66mM

Enzyme Activities

Ten New Terms

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