Transcript Microbial Genetics

Microbial
Genetics
Central Dogma
Regulation of Gene Expression
Genetic Exchange
Precursors and “Building Blocks” of
Nucleic Acids
Nucleotides
polymerize to a
nucleic acid by
phosphodiester
bonds.
=O
An organic ester:
R-C-O-C-R
=O
A phosphodiester:
=O
R-C-O-P-O-C-R
Base Pair
Complementation
Pyrimidines: C & T (U in RNA)
Purines: G &A
G to C by 3 H-bonds
A to T (U) by 2 H-bonds
High G+C content of DNA
means more H-bonds;
requires more energy to
separate (denature) duplex.
RNA Structure:
Single Strand with Self Complementation
hairpin loop structure
(see transcription
terminator region )
5’-P
3’-OH
A G
C
T
T
C
A
G
C
A
T
G
5’-P
3’-OH
3’-OH
5’-P
DNA Structure:
Complementation of
Two Antiparallel
Strands
Forms into a Double
Helix Conformation
DNA
Replication
It is semi-conservative;
half from the “parent” half
newly synthesized.
It’s initiated at a replication
fork; DNA must be
unwound and unbound
into two single strands.
Parent strands become
the templates for the new
(complementary) strands.
DNA Polymerase
Reaction
It must add to an existing
nucleic acid (RNA or DNA).
Adds dNTP to the 3’-OH end
of a new strand; it grows in
the 5’→ 3’ direction.
DNA-Poly III has
“Proofreading” capability; can
catch and fix mistakes most
of the time by 3’→ 5’
exonuclease activity.
Enzyme Functions in Replication
DNA Polymerase III
This is a simple cartoon;
exaggerated to illustrate function.
Primosome (makes RNA primer)
Ligase
(connects Ofragments)
DNA Polymerase I (also has RNase, 5’ → 3’
exonuclease, activity to remove RNA primer)
Replisome
Two DNA Polymerase III stay together in a complex called
the replisome.
This requires the lagging strand to loop back onto its
strands DNA Polymerase III.
When the lagging strand DNA Polymerase reaches the
previous Okazaki Fragment it drops off and then grabs the
next RNA primer.
Prokaryote
Chromosome
Replication
(OriC)
Chromosome is of circular DNA, and
supercoiled (extra twists in helix knotsup or condenses DNA when not
replicating).
Plasmids = extrachromosomal
element; circular; 100-1000 times
smaller than chromosome. Also
supercoils when not replicating.
Plasmid DNA replication like that of
chromosome or by rolling circle
replication (more later in discussing
bacterial conjugation, “sex”).
Termination
(ter)
Replication Initiation
• Initiator proteins (IP; about 30) bind at repetitive
sequences within the OriC site.
• DNA winds around IP-complex; induces separation of
strand at adjacent AT rich site.
• Single strand binding proteins & helicases attach.
• Formation of two replication forks that open in opposite
directions.
~30