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E. coli happy Excess of amino acids. E. coli moderately happy Interconversion of amino acids. Too much alanine. Strategy. glucose. Too little phenylalanine. E. coli moderately happy Interconversion of amino acids. Too much alanine. Transaminase. Oxaloacetic acid (OAA). Aspartate.
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E. colimoderately happyInterconversion of amino acids Too much alanine Strategy glucose Too little phenylalanine
E. colimoderately happyInterconversion of amino acids Too much alanine Transaminase Oxaloacetic acid (OAA) Aspartate Transaminase Enough phenylalanine Aspartate
E. coliconcernedFormation of amino acids de novo Alpha-keto acidsbut no amino acids Oxaloacetic acid (OAA) nitrate
E. coliconcernedFormation of amino acids de novo Nitrogen Starvation nitrate Nitrate reductase α-ketoglutarate ammonia Glutamine synthetase glutamine glutamate Glutamine synthetase glutamate
E. coliconcernedFormation of amino acids de novo Nitrogen Starvation Glutamine synthetase Nitrate reductase Presence of amino acid
E. coliconcernedFormation of amino acids de novo Nitrogen Starvation Environmental signal Intracellularsignal Sensation of nitrogen starvation Presence of nitrate Presence of histidine Response Glutamine synthetase Nitrate reductase Histidine degradation
Control of glutamine synthetase Active glutamine synthetase P transcription translation gln Posttranscriptionalcontrol ATP Adenylylation Transcriptionalcontrol Inactive glutamine synthetase
Control of glutamine synthetase Nitrogen Starvation pppGpp! ??? Translation of signal Stringent response evoked by N starvation …but wrong response! If not pppGpp then what?
Control of glutamine synthetaseAutogenous regulation? Effect of Gln mutants on nitrogen control (Ntr) Mutations mapping to gln affect global N control GS need not be active to exert control
Control of glutamine synthetaseMutations outside of glnA glnA X Y Maybe all those glnA mutations were really in X or Y??? Yes and no So GS STILL regulates itself? Maybe not.
Control of glutamine synthetaseMutations outside of glnA P glnA X Y transcription
Control of glutamine synthetaseMutations outside of glnA P glnA X Y stop transcription Polar effect of nonsense mutations Plausible,… ...but is it true?
Control of glutamine synthetaseAre “regulatory” glnA merely polar?How to tell? P NtrC glnA X Y P NtrC glnA X Y Complementation: If not polar, then yes If polar, then no
Control of glutamine synthetaseCurrent understanding of glnA operon P glnA ntrB ntrC N-regulated gene expression P Phosp’nof NtrC NtrB NtrC UMP Uridyl’nof PII Control of NtrB PII (GlnB) Uridylyltransferase (UT)(GlnD)
E. coliconcernedFormation of amino acids de novo Nitrogen Starvation Environmental signal ??? Intracellularsignal Sensation of nitrogen starvation Presence of nitrate Presence of histidine Response Glutamine synthetase Nitrate reductase Histidine degradation
Control of glutamine synthetaseHow is nitrogen status sensed? Uridylyltransferase (UT) UMP - + Active PII Inactive PII Uridylyltransferase (UT) glutamine α-ketoglutarate
E. coliconcernedFormation of amino acids de novo Nitrogen Starvation nitrate Nitrate reductase α-ketoglutarate ammonia Glutamine synthetase glutamine glutamate Glutamine synthetase glutamate