Lecture 9 1
2 Component Systems
Generally, bacteria live in environments that undergo
DRASTIC CHANGE (resources and conditions)
To survive, they must respond rapidly, through:
1. adaption 2. Movement (fight or flight)
Therefore, bacteria have evolved mechanisms to sense
external conditions & transfer this information to cellular
sites where an appropriate response is exerted.
> altered patterns of gene expression
This SIGNAL TRANSDUCTION generally involves the
transfer of phosphoryl groups between proteins.
Many of these signals are sensed and transmitted to the Tn
apparatus by PAIRS of proteins, called Called a 2 component
Sensor Kinases (SK) & Response Regulators (RR)
> 1000 pairs of
cognate pairs of Lecture 9 2
SK + RR. Seen NtrB/C already. Also, FixL/FixJ. NtrB
phophorylates NtrC. NtrB is SK. NtrC is RR.
Autophsorylation of sensor kinase > phosphorylation of
RR > altered confromation > DNA binding by (1) RR itself
or (2) other proteins > altered Tn.
NtrB is an example of a bi-functional kinase/phosphates. (needs to by able to chut off signal
when not needed. Either SK is bi-functional or there is another phophotase involved. Lecture 9 3
Various regulatory mechanisms (some of which act
through HPK / SK) regulate intracellular concentration of
RR in response to stimuli.
When the phosphorylated form of RR (RR ) reaches a
critical level, the DNA is bound, leading to RNAP
recruitment (activation) or interference (repression).
The physiological adaption is regulated via control of the
concentration of the phophorylated for RR-P of the RR. Lecture 9 4
Autophosphorylation Phosphorylation of RR
N-terminal conserved domain ~125 AA with 30% identity
between proteins and several invariant AA’s.
This is the phosphate Receiver Domain – 3 conserved
Asp / 1 Lys
The aspartates form an acidic pocket; lysine protrudes into
the pocket to stabilize the phosphoryl group.
C-terminal conserved domain = OUTPUT Domain (or
These interact with the transcription complex (RNAP).
They mediate Tn Regulation by either:
1. Altered DNA binding (by themselves)
2. Altered RNAP binding. Lecture 9 5
The change in the activity of the OUTPUT domain is
brought about through reversible transfer of phosphoryl
groups to invariant aspartates at receiver domains.
3 distinct sub-families of RRs
Based on structural and functional organization of the
C-term DNA-binding HTH = 150 AA – linked to receiver
When receiver domain is phosphorylated, a conformational
change causes STRONG DNA binding by output domain.
Dephosphorylation causes DNA-binding interference. Lecture 9 6
Dephosphorylation causes DNA-binding interference.
OmpR-like 2 component systems act at σ70 promoters.
Tn can be either activated or repressed by OmpR-like 2
component systems, depending upon where RR binds.
For some of these 2C systems, they bind DNA and make
direct contact with αCTD of RNAP.
Conserved C-terminal DNA-binding HTH = 100 AA;
different sequence from OmpR type
Act in combination with σ70 RNAP / promoters
1. many homologs of FixJ RR’s have an output domain,
but no reciever domain.
2. Other FixJ RR’s have receiver domains, but no output
domain (no gene regulation).
e.g. – CheY: phosphorylated CheY interacts with flagellar
switch proteins to cause reversals in flagellar rotation.
This is protein-protein interaction. Not DNA interaction. Lecture 9 7
NtrC - 3 separate domains
• N term – Reciever domain
• C term – Dna-binding HTH
• Central domain = ATPase – similar to ATP binding
Unlike OmpR & FixJ which act by improving RNAP-DNA
interaction (RNAP recruitment), NtrC-like RR’s facilitate
This RXN involves the ATPase function.
ATPase activity is dependant on prior phosphorylation of
the reciever domain leading to formation of tetramers and
aggregates of Ntr-C like proteins Lecture 9 8
In the de-phosphorylated state these RR’s exist as dimers
and have no activity.
Transfer of the phosphoryl group is maintained by HPK.
However, for each RR, a COGNATE HPK can be assigned.
They are named HPK because the source of the
phosphoryl group is a phospho histadine AA.
Modular structure; conserved 250 AA’s; 20% ID; 5 sub-
domains: H box histidine is the active site which catalyzes
transfer of γP from ATP to histidine AA. ATP binding site is
in the G box.
ATP binding site in G box Lecture 9 9
Usually HPKs exist as dimers; Phosphorylation occurs