bacterial chromosome
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Bacterial Chromosome•Free-living bacteria need genetic information to synthesize proteins
for executing vital functions.
•Most bacteria have a single chromosome with DNA that is about
2Mbp (mega base pairs) long (1Mbp=1X106
base pairs), but the DNAcontent of different species varies from 0.58 to greater than 9 Mbp of
DNA.
•In contrast to the linear chromosomes found in eukaryotic cells, the
strains of bacteria initially studied were found to have single,
covalently closed, circular chromosomes.
•The circularity of the bacterial chromosome was elegantlydemonstrated by electron microscopy in both Gram negative
bacteria (such as Escherichia coli ) and Gram positive bacteria (such
as Bacillus subtilis ).
•Bacterial plasmids were also shown to be circular.
•In fact, the experiments were so beautiful and the evidence was so
convincing that the idea that bacterial chromosomes are circular andeukaryotic chromosomes are linear was quickly accepted as a
definitive distinction between prokaryotic and eukaryotic cells.
•However, like most other distinctions between prokaryotic and
eukaryotic cells, it is now clear that this dichotomy is incorrect.
•Not all bacteria have a single circular chromosome: some bacteria
have multiple circular chromosomes, and many bacteria have linear
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chromosomes and linear plasmids.
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•Some bacteria have multiple chromosomes.
•The first convincing evidence that some bacteria have multiple
chromosomes came from studies on Rhodobacter sphaeroides .
•Both molecular and genetic studies clearly demonstrated that R.
sphaeroides has two large circular chromosomes.
•One of the chromosomes is 3.0 Mb and the other is 0.9 Mb.
•Leptospira also has two chromosomes of 4.4 and 4.6 Mbp.
•The largest bacterial genome yet analysed is that of Myxococcus
xanthus, with 9.2Mbp.
•However, the best studied organism in nature is E. coli , which has a
4.6-Mbp chromosome with 4288 genes for proteins, seven operons forribosomal ribonucleic acids (RNAs), and 86 genes for transfer RNAs.
•Some bacteria have linear chromosomes also.
•The first published evidence for linear chromosomes was in 1979,
but because the techniques used at that time were limited and
because the dogma that all bacterial chromosomes are circular was
so entrenched, few people believed that linear chromosomes andplasmids occured in bacteria until 1989.
•Borrelia have linear chromosomes and most strains contain both
linear and circular plasmids; most of the bacteria in the genus
Streptomyces have linear chromosomes and plasmids and some have
circular plasmids as well.
•In addition, in some cases there may be a dynamic equlibrium
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between linear and circular forms of a DNA molecule.
•There is some evidence that linearization may be due to integration
of a linear phage genome into the circular DNA molecule.
•On a macroscopic scale, bacterial chromosomes are either circular
or linear.
•Circular chromosomes are most common, at least among the best-
studied bacteria.
•However, the causative agent of Lyme disease, Borrelia burgdorphei ,
has a 2-Mb linear chromosome plus 12 different linear plasmids.
•Eukaryotic chromosomes are invariably linear, and they have two
ends, each a telomere, and a organized region, the centromere whichallows the chromosome to attach to cellular machinery that moves it
to the proper place during cell division.
•The ends of linear DNA molecules (called telomeres) pose two
problems that do not apply to circular DNA molecules.
•First, since free double-stranded DNA ends are very sensitive to
degradation by intracellular nucleases, there must be a mechanismto protect the ends.
•Second, the ends of linear DNA molecules must have a special
mechanism for DNA replication.
•These problems are solved by features of the telomeres.
•Two different types of telomeres have been observed in bacteria:
hairpin telomeres and invertron telomeres.
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•Linear DNA molecules in bacteria are protected by both types of
telomeres: palindromic hairpin loops are protected by the lack of
free double-stranded ends, and invertron telomeres are protected by
proteins that bind to the 5'-ends.
•Both of these mechanisms are also used by some phage, eukaryotic
viruses, and eukaryotic plasmids.
•Telomeres also solve the problem of DNA replication differently.
•Invertron telomeres have a protein covalently attached to the 5'
ends of the DNA molecule (called the 5'-terminal protein or TP).
•DNA polymerase interacts with the TP at the telomere and catalyzesthe formation of a covalent bond between the TP and a dNTP.
•The dNTP bound to the TP has a free 3'-OH group which acts as the
primer for chain elongation.
•Replication of hairpin telomeres is less well understood.
•Apparently multiple hairpin sequences can pair to form concatemers
that are replication intermediates.
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•One critical facet of chromosome structure is that DNA is a
plectonemic helix, which means that two helical strands entwine
about each other.
•For duplex DNA the two antiparallel strands are interwound once for
every 10 bp.
•Because of this wound configuration, biochemical transactions that
involve strand separation require chromosome movement (spin)
about DNA’s long axis.
•The processes ofDNA replication, recombination and transcription all
require DNA rotation, and during DNA synthesis the rotation speed
approaches 6000 rpm.
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•Chromosomal DNA molecules are
very long and thin, so DNA must
fold many times to fit within the
confines of a bacterial cell.
•How does DNA twisting transpire
without chromosomes getting
tangled up?
•The problem was important
enough that Watson and Crick
considered a paranemic helix,
which is a pair of coils that layside by side without interwinding.
•Strands of a paranemic helix can
separate without rotation.
•None the less DNA is
plectonemic, and keeping
chromosomes untangled requiresa special class of enzymes called
topoisomerases.
•In the dimensions of B-form DNA,
E. coli is a sphere that is 6 kbp
long (8 nm) and 4 kbp wide (2 nm),
so the 4.6-Mbp chromosome must
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be folded many times to fit within a cell.
•Negative supercoiling forces DNA into an interwound configuration.
•First, the DNA molecule doubles back on itself so that length is
halved relative to that of the linear form.
•Second, supercoiled branches are dynamic so that opposing DNA
sites in the interwound network are constantly changing.
•A protein bound to DNA in one supercoiling domain interacts more
than 100 times more frequently with other proteins in the same
domain than it does with proteins bound to a different domain.
•When DNA is liberated from cells by breaking the peptidoglycan coat,
chromosomes form bundled loops that represent domains.•Such preparations (called nucleoids) behave as discrete bodies.
•Many reactions ofthe chromosome require the formation of intricate
DNA – protein machines to replicate, transcribe or recombine DNA at
specific sequences.
•A group of ‘chromosome-associated’ proteins assists in the
formation of these complexes by shaping DNA.•These proteins are sometimes described as histone-like, although
they share no structural similarity with the eukaryotic histones.
•Chromosome-associated proteins include HU, H-NS, intregation host
factor (IHF) and factor for inversion stimulation (FIS).
•HU is encoded by two genes, hupA and hupB, which are closely
related to each other and to the genes encoding the two IHF
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subunits.
•HU is the most abundant doublestranded DNA-binding protein in E.
coli, although it also binds RNA.
•Although HU shows a preference for structurally ‘bendable’
sequences, it binds DNA relatively nonspecifically, forming
complexes that stabilize negative supercoils in double-stranded DNA.
•H-NS is a second relatively nonspecific DNA-binding protein that
forms dimers, tetramers and possibly higher order associations when
bound to DNA.
•Like HU, H-NS constrains negative supercoils.
•
H-NS binds ‘bendable’ DNA, which include AT-rich sequences oftenfound near bacterial promoters; H-NS also modulates the
transcription of numerous operons in E. coli and of lysogenic phages.
•FIS protein forms homodimers and binds DNA at specific sites.
•FIS was discovered nearly simultaneously in two related site-specific
recombination systems: the Hin inversion reaction of Salmonella
typhimurium and the Gin inversion reaction ofphage Mu.•FIS concentration rises in early log phase when the protein has
regulatory roles in replication initiation at oriC and transcription of
ribosomal RNA operons, and then FIS abundance declines in
stationary phase.
•IHF protein is composed oftwo subunits (encoded by the himA and
hip genes) that are about 100 amino acids long and which make
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stable heterodimers.
•IHF protein binds to a consensus sequence that was first discovered
in phage λ, where it stimulates insertional and excisional
recombination in vitro by a factor of 104.
•IHF bends DNA severely (more than 90º); such bends are critical for
site specific recombination in λ, for repression and activation of
phage Mu, and for repression – activation in a number of cellular
operons, including ilv, oxyR and nifA.
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Supercoiling
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The diverse group of prokaryotes share even more common features with the
eukaryotes.
•The circular genomes of mitochondrial and chloroplast are a notable exception to the
rule that eukaryotic chromosomes are linear. However, this nicely fit into the dichotomy
that eukaryotic chromosomes are linear and bacterial chromosomes are circular
because these organelles seem to have evolved from entrapped bacteria.
•Other examples include the presence of introns, and poly-A tails on mRNA.•This genus includes B. burgdorferi, the causative agent of Lyme disease.
1.Streptomyces make a wide variety of useful antibiotics, including streptomycin.
2.For example, linear DNA was precipitated in the most commonly used procedures for
purifying bacterial plasmids, and the procedures for purifying chromosomal DNA relied
upon the differential binding of ethidium bromide to "sheared DNA fragments" compared
to circular DNA.
3.It is not intuitively obvious how the ends of a linear DNA molecule could be completely
replicated. All known DNA polymerases require a pre-existing primer for initiation of DNA
replication. The primer is usually a short RNA molecule with a free 3'-OH group that can
be extended by DNA polymerase. If a linear DNA molecule was primed at one end, DNA
synthesis could continue to the other end. However, once the primer is removed, the DNA
corresponding to the primer could not be replicated.
The telomers at the end of chromosomes of most eukaryotic cells are replicated by a
different mechanism: most telomeres are short GC-rich repeats that are added in a 5' to
3' direction by the enzyme telomerase