2 component regulatory systems

whooshdisguisingBiotechnology

Dec 14, 2012 (4 years and 7 months ago)

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2 component regulatory systems


Maltose=effector, BUT if signal not DIRECTLY involved, but needs
to be transmitted and changed = signal transduction


Sensor protein=


kinase, phosphorylates compounds,


membrane associated


Phosphoryl group transmitted to another regulator IN the cell


Often a DNA binding protein involved in transcription


Many examples, N
-
fixation, sporulation,chemotaxis


Chemotaxis


Attractants decrease rate of autophosphorylation


Repellant increased autophosphorylation


CheA
-
CheW=transducer


CheY controls switch


cheY
-
P tumbles, CCW
-
CW


CheB phosporylated by CheA
-
P, but slower response than
CheY
-
P


CheB involved methylation


Fully methylated = best for repellants


cheB
-
P demethylates, occurs when attractants High


Degree of methylation regulates attraction/repulsion


Chemotaxis

Genetic exchange: transformation

Genetic exchange: plasmid transfer


Small, usually circular, independently replicating DNA
molecules


Generally, G
-

plasmids replicate as does chromosomal DNA, G
+

plasmids by “rolling circle” replication


Genes of replication control, timing initiation on plasmid (
ori
)


Some plasmids integrate (F
+
, Hfr)


Most are double
-
stranded


About 1
-

100kb


Code for:


R
-
factors (R
-
plasmids) : antibiotic resistance, heavy metal
resistance


Virulence plasmids : adhesins, hemolytic factors, toxin, Ti,
bacteriocins


Degradation, tol, nah,


Plasmid copy #


Compatibility (
inc
)




Genetic exchange: plasmid transfer


Small, usually circular, independently replicating DNA
molecules


Generally, G
-

plasmids replicate as does chromosomal DNA, G
+

plasmids by “rolling circle” replication


Genes of replication control, timing initiation on plasmid (
ori
)


Some plasmids integrate (F
+
, Hfr)


Most are double
-
stranded


About 1
-

100kb


Code for:


R
-
factors (R
-
plasmids) : antibiotic resistance, heavy metal
resistance


Virulence plasmids : adhesins, hemolytic factors, toxin, Ti,
bacteriocins


Degradation, tol, nah,


Plasmid copy #


Compatibility (
inc
)




Avery Experiment

Transformation


Competence
: ability to be transformed


Steps in transformation


DNA binding and uptake (SS or DS, depending
on species)


Integration (recA)


Competence may be induced by
electroporation, Ca


Mechanism of transformation

Transduction


Generalized


low frequency


Specialized


high frequency:requires
specific integration

Generalized transduction

Specialized transduction

Specialized transduction

Genetic exchange: conjugation

Conjugation: early

Cell
-
surface
structure

Conjugation: middle

Conjugation: late

Genetic Engineering
-
basics


Basic steps in cloning


Restriction
-
modification enzymes


“shot gun” or PCR


Ligase


Recombination Plasmids (or phage)=
vectors


Expression vectors


Selection of clones


Looking for a clone with a specific gene


Probes (DNA, RNA or antibody)


Wave of the future: DNA chips or “microarrays”, BAC
libraries, automated sequencing etc


Genomics


Bioinformatics


Harvesting genes for biotech (Diversa)


Recent Science article


Comparing gene families