MICR3240 - Nitrogen utilization

Strategy 1 

millimolar range,using glutamate dehydrogenase, used in high ammonium cases where less energy is required

Strategy 2

micromolar range, glutamine synthetase, works better in low ammonium conc. at the expense of energy

GS allosterically inhibited by

many end-products of N assimilation

AT adenylates the Tyr of GS the consequence of this is

that it reduces the GS activity, can be a spectrum of how many units get adenylated

Adenylation causes a switch from

Mn to Mg causing feedback inhibition 

Uridylation and
deuridylation of PII are catalyzed by

UMP

high gln inhibits and activate

uridyl transferase and uridyl hydrolase

When [glutamine]/[alpha-ketoglutarate] ratio is high

PII not uridylated, PII stimulates adenylation of GC, adenylated Gs is less active and sensitive to inhibition by products of N-assimilation 

When [glutamine]/[alpha-ketoglutarate] ratio is low

PII is uridylated to PII-UMP, PII-UMP cannot stimulate adenylation of GS, unadenylyated GS is highly active

What regulates gene expression of glnA

ntrB and ntrC

on the glnA sigma 70 is a and sigma 54 is a 

weak promoter, strong promoter 

Role of Ntr-C

as an activator, when phos. It binds upstream of promoter away from sigma 54, to be able to interact with rna polymerase the dna has to bend putting the activator in contact with rna polymerase to be able to hold sigma 54 long enough to start the transcription process - open complex formation

What helps NtrC-P with the open complex formation

IHF

Transcription of glnA from the sigma54 promoter is regulated by a two component system

NtrB sensor kinase and NtrC response regulator

Sensor kinase 

NtrB receives a signal under nitrogen
limiting condition and responds by autophosphorylating HIS

NtrB can transfer phos. to

response regulator NtrC

Ammonia Excess ([Gln]/[aKG] high)

PII is not uridylylated, phosphatase activity of NtrB is enhanced by association with PII

Ammonia Limitation ([Gln]/[aKG] low

PII is uridylylated by UTase and it rrleases NtrB phos. NtrC, Open complex formation/transcription from glnAp2 is
activated, Transcription from glnAp2 expresses all 3 genes and the level of GS rises 30-fold

Free living heterotrophs 

Azotobacter, bacillus 

Plant symbionts or plant associated bacteria 

rhizobium

Free living, facultatively aerobic diazotroph

use N2 only if O2 is low, energy supplies are abundant and better N supply is available

Symbiotic, facultatively aerobic diazotrophs

rely on the microaerophilic environment within plant root nodules and C/energy supplied by the plant

Anaerobic diazotrophs

have novel mechanisms for active energy metabolism without O2 

Photosynthetic, aerobic diazotrophic cyanobacteria

segregate their oxygenic photosynthetic apparatus from their nitrogen fixing one in space