1) Prokaryotic chromosomes are not enclosed in a nucleus, and their DNA is dispersed within the cell. Much of what is known about DNA structure comes from studying prokaryotes. 2) Prokaryotes have a single circular chromosome that must be tightly packed to fit in the cell. In E. coli, its chromosome is condensed through supercoiling and organized into loops attached to the cell membrane. 3) Nucleoid-associated proteins are important for condensing and organizing the bacterial chromosome through bridging, wrapping and constraining DNA supercoils. The most extensively studied in E. coli are HU, IHF, H-NS, and Fis.

1. PROKARYOTIC CHROMOSOME
STRUCTURE
Dr. Esther Shoba R
Assistant Professor
Kristu Jayanti College

5.  The term “prokaryote” means “primitive nucleus”. Cell in
prokaryotes have no nucleus. The prokaryotic chromosome is
dispersed within the cell and is not enclosed by a separate
membrane.
 Much of the information about the structure of DNA has come
from studies of prokaryotes, because they are less complex
(genetically and biochemically) than eukaryotes.
 Prokaryotes are monoploid = they have only one set of genes
(one copy of the genome).
 In most viruses and prokaryotes, the single set of genes is
stored in a single chromosome (single molecule either RNA or
DNA).
 The smallest known RNA viruses have only three genes.
 The smallest known DNA viruses have only 9 to 11 genes.

6.  The bacterial chromosome must be tightly packed to fit into
the small volume of the bacterial cell. The contour length of
the circular DNA molecule present in the chromosome of the
bacterium Escherichia coli is about 1500 µm. Because an E.
coli cell has a diameter of only 1 to 2 µm, the large DNA
molecule must exist in a highly condensed (folded or coiled)
configuration.
 Compacting the DNA involves supercoiling, or further twisting
the twisted chromosome.
 The chromosome's fifty or so DNA domains are held together
by a scaffold of RNA and protein, and the entire nucleoid is
attached to the cell membrane.
 This membrane attachment aids in the segregation of the
chromosomes after they replicate in preparation for cell
division.
 Bacteria lack the histone proteins that are found bound to
DNA and that form a nucleosoms of eukaryotic chromosomes.

7.  DNA molecule in an E. coli chromosome is organized into 50 – 100
domains or loops.
 Replication of the circular chromosome begins at a single point,
called OriC, and proceeds in both directions around circle, until the
two replication forks meet up.
 The results is two identical loops. Replication takes approximately
forty minutes.
 In 1997 F Blattner and colleagues published the sequence of
4,639,221 base pairs of the K-12 laboratory strain. E. coli is
estimated to have 4,279 genes.
 Many sets of genes on the E. coli chromosome are organized into
operons.
 An operon is a set of functionally related genes that are controlled by
a single promoter and that are all transcribed at the same time.
 It is also quite common for bacterial species to possess extra
chromosomal genetic elements called plasmids. These are small,
circular DNA molecules which, when present, vary in umber from
one to about thirty identical copies per cell.
 Plasmids include the fertility factor, as well as plasmids that carry
drug-resistance genes.

13. WHAT HELPS CONDENSATION AND ORGANISATION
• NUCLEOID ASSOCIATED PROTEINS
• NAPs are highly abundant and constitute a significant
proportion of the protein component of nucleoid
• NAPs induce and stabilize bends in DNA, thus aid in DNA
condensation by reducing the persistence length. NAPs
condense DNA by bridging, wrapping, and bunching that could
occur between nearby DNA segments or distant DNA segments
of the chromosome.

14. • Another mechanism by which NAPs participate in
chromosome compaction is by constraining negative
supercoils in DNA thus contributing to the topological
organization of the chromosome.
• There are at least 12 NAPs identified in E. coli, the most
extensively studied of which are HU, IHF, H-NS, and Fis.
Their abundance and DNA binding properties and effect on
DNA condensation and organization are summarized in the
tables below

18. HU
• Histone-like protein from E. coli strain U93 (HU) is an
evolutionarily conserved protein in bacteria. HU exists in E.
coli as homo- and heterodimers of two subunits HUα and HUβ
sharing 69% amino acid identity.
• HU is a non-sequence specific DNA binding protein. It binds
with low-affinity to any linear DNA. However, it preferentially
binds with high-affinity to a structurally distorted DNA.
• Examples of distorted DNA substrates include cruciform
DNA, bulged DNA, dsDNA containing a single-stranded
break such as nicks, gaps, or forks.

19. • Furthermore, HU specifically binds and stabilizes a protein-
mediated DNA loop.
• In the structurally specific DNA binding mode, HU recognizes a
common structural motif defined by bends or kinks created by
distortion, whereas it binds to a linear DNA by locking the
phosphate backbone.
• While the high-affinity structurally-specific binding is required
for specialized functions of HU such as site-specific
recombination, DNA repair, DNA replication initiation, and
gene regulation, it appears that the low-affinity general binding
is involved in DNA condensation
• one HU dimer every ~150 bp of the chromosomal DNA based
on the estimated abundance of 30,000 HU dimers per cell
(4600000 bp /30,000)

20. IHF –INTEGRATION HOST FACTOR
• Integration host factor (IHF) is structurally almost identical to
HUbut behaves differently from HU in many aspects.
• Unlike HU, which preferentially binds to a structural motif
regardless of the sequence, IHF preferentially binds to a
specific DNA sequence even though the specificity arises
through the sequence-dependent DNA structure and
deformability.
• The specific binding of IHF at cognate sites bends DNA sharply
by >160-degree.

21. An occurrence of the cognate sequence motif is about 3000 in the
E. coli genome.
The estimated abundance of IHF in the growth phase is about
6000 dimers per cell.
Assuming that one IHF dimer binds to a single motif and nucleoid
contains more than one genome equivalent during the exponential
growth phase, most of the IHF molecules would occupy specific
sites in the genome and likely only condense DNA by inducing
sharp bending.