Download I. virAL CHROMOSOMES

Chromosomes and Chromosomal Folding
I.
VIRAL CHROMOSOMES
A. Viral genomes may be either RNA or DNA
1. RNA and DNA are never both found together in a virion
B. Viral genomes may be circular (like prokaryotes) or linear (like eukaryotes)
C. Viral genomes may be single-stranded or double stranded
D. When packaged into the virion the genome is totally inactive until it is delivered to
the next host cell
II.PROKARYOTIC
CHROMOSOMES
A. Characteristics
1. Located in the nucleoid region
a) It is not surrounded by a nuclear membrane
2. Single double-stranded covalently-closed circular DNA
a) ds cccDNA
3. 1300 m in length
a) Typical prokaryotic cell is about 1 m in length
b) Center of the chromosome is attached to the plasma membrane
(1) This helps partition the chromosomes during cell division
4. It is negatively supercoiled
B. Detection of cccDNA
1. Alkaline sedimentation ultracentrifugation
a) DNA is denatured above pH 11.3
(1) Linear DNA
(a) Will spearate into two strands
(b) S will increase 30%
(2) cccDNA
(a) Will result in a tangle mess
(b) S will increase 300%
(3) Nicked circular DNA
(a) Will result in one linear and one circular DNA molecule
(b) S will increase 44%
2. Ethidium bromide CsCl-density ultracentrifugation
a) As ethidium bromide intercalates between bases, it unwinds the DNA, thus
decreasing its density
b) Linear DNA
(1) Will unwind the most
(2) Will be least dense
c) cccDNA
(1) Will unwind the least
(2) Be most dense
d) Nicked circular DNA
(1) Will have intermediate density
3. Electrophoresis
a) Supercoiled molecules migrate faster than non-supercoiled molecules
C. The DNA is arranged in a series of loops
1. Each loop is supercoiled
a) The loops are attached to proteins
(1) Attachment to proteins prevent rotations past loops
(2) Proteins are basic (positively charged) allowing them to interact with the
negatively charged DNA
2. The number of loops can be determined by the number of cuts needed to
remove all supercoiling
a) Removing all supercoiling should reduce S value by 30%
b) Single-stranded breaks remove supercoiling from a loop
(1) It takes 45 - 50 breaks needed to obtain this with E. coli chromosome
c) This corresponds to the number of DNA gyrase binding sites
(1) DNA gyrases introduces negative supercoiling
III.MITOCHONDRIAL
AND CHLOROPLAST CHROMOSOMES
A. These organelles are largely inherited from the female parent
1. Transmission of diseases related to mutations in these organelles are inherited
maternally
B. The genome is similar to prokaryotic chromosomes
1. ds cccDNA packaged into loops
2. Lack histones of nuclear chromosomes
C. Size and numbers
1. The genome is about 16,600 base pairs in humans
2. There is approximately 5 to 10 copies of the genome per organelle
D. Replication
1. Organelle DNA is replicated by enzymes encoded in the nucleus
IV.EUKARYOTIC
CHROMOSOMES
A. General
1. If proteins are removed from a eukaryotic chromosome it would be about 2 nm
thick by 12,000,000 nm long
2. Terminology
a) Twisting number
(1) Total number of turns of the helix
(2) In non-supercoiled molecules, this would equal the total number of base pairs
divided by the number of base pairs per turn of the helix
b) Writhe
(1) Number of nodes
(2) W = 0 for nicked circles
B.
C.
D.
E.
c) Linking number
(1) The number of times the two strands cross each other
(2) T = L in non-supercoiled molecules
d) L = W + T
(1) If T decreases, either L will decrease or W will increase
(2) The strain of underwiding can be accommodated by negative supercoiling
(a) Single stranded region increases with temperature
(b) Negative supercoiling decreases with temperature
(3) Breathing is important for recombination, initiation of replication, and initiation
of transcription
Histones
1. Histones are basic proteins containing large amounts of the cationic amino
acids lysine and arginine arranged mostly in  helices
a) The positive charges allow the proteins to interact with the negatively
charged DNA
b) The alpha helices fit into the major groove of DNA
2. 5 classes of histones
a) H1, H2A, H2B, H3, and H4
b) Highly conserved
(1) There is only a 2 amino acid differences between cows and peas which
diverged 109 years ago
Nucleosomes
1. Nucleosomes are DNA wrapped around an octamer of histone proteins
a) The DNA associated with the histones is referred to as the core DNA
(1) H2A, H2B, H3 & H4
(2) Core DNA is about 146 base pairs in length
b) The space between nucleosomes is referred to as linker DNA
(1) Length of linker DNA varies between tissues and organisms
(2) Linker DNA is associated with H1
c) The DNA associated with histones has a 'bead on a string' appearance
(1) It is about 11 nm in diameter
Solenoids
1. H1 histone will link the nucleosomes into a solenoid structure
a) The solenoid is about 30 nm in diameter
b) This is the state of DNA in a resting nucleus
Scaffolding
1. Chromosomes can compact 100-fold more by scaffolding with non-histone
proteins
a) The scaffolded chromosome is around 700 nm thick
b) This is the state of DNA during mitosis and meiosis