Supercoiling, bacterial

Bacterial as well as eukaryotic chromosomes contain too much DNA to fit easily into a cell. Therefore, the DNA must be condensed (compacted) to fit into the cell or nucleus. This is accomplished by supercoiling the DNA into a highly condensed form. When relaxed circular DNA is twisted in the direction that the helix turns, the DNA becomes positively supercoiled, if it is twisted in the opposite direction, it is called negatively supercoiled. Bacterial DNA is normally found in a negatively supercoiled state. Supercoiling reactions are catalyzed by topoisomerases. 

Gyrase is another term for bacterial topoisomerase II. The enzyme consists of two A and two B subunits and is responsible for the negative supercoiling of the bacterial DNA. Negative supercoiling makes the bacterial DNA more compact and also more readily accessible to enzymes that cause duplication and transcription of the DNA to RNA. 

Supercoiling with bacterial DNA gives a largely open, extended, and narrow, rather than a compacted, multibranched structure. By comparison, the DNA in eukaryotic cells is present... 

Fluoroquinolones are inhibitors of bacterial topoisomerase 11 (DNA gyrase). This enzyme reduces the supercoiling of DNA in order to allow separation of the two strands of DNA that are required for replication and transcription. In the first stage, topoisomerase II cleaves the two strands of DNA, after repairing it, and restores the supercoiling of DNA. ... 

Correct answer = C. Fluoroquinolones, such as ciprofloxacin, inhibit bacterial DNA gyrase—a type II DNA topoisomerase. This enzyme catalyzes the transient breaking and rejoining of the phosphodiester bonds of the DNA backbone, to allow the removal of positive supercoils during DNA replication. The other enzyme activities mentioned are not affected. Primase synthesizes RNA primers, helicase breaks hydrogen bonds in front of the replication fork, DNA polymerase I removes RNA primers, and DNA igase joins Okazaki fragments. 

Isolated "naked" bacterial DNA, from which proteins have been removed, is supercoiled. DNA in the bacterial chromosome is also supercoiled. When naked DNA is nicked, its supercoiling is abolished. In contrast nicking the chromosomal DNA does not abolish its supercoiling. Explain. 

The DNA of a bacterial cell, such as Escherichia coli, is a circular double-stranded molecule often referred to as the bacterial chromosome. In E. coli this DNA molecule contains 4.6 million base pairs. The circular DNA is packaged into a region of the cell called the nucleoid (see Topic Al) where it is organized into 50 or so loops or domains that are bound to a central protein scaffold, attached to the cell membrane. Fig. la illustrates this organization, although only six loops are shown for clarity. Within this structure, the DNA is actually not a circular double-stranded DNA molecule such as that shown in Fig. lb but is negatively supercoiled, that is, it is twisted upon itself (Fig. lc) and is also complexed with several DNA-binding proteins, the most common of which are proteins HU, HLP-1 and H-NS. These are histone-like proteins (see below for a description of histones). 

Fig. 1. (a) The association of circular bacterial DNA with a protein scaffold (b) a circular double-stranded DNA molecule (c) supercoiled DNA. 

One of the most exciting biological discoveries is the recognition of DNA as a double helix (Watson and Crick, 1953) of two antiparallel polynucleotide chains with the base pairings between A and T, and between G and C (Watson and Crick s DNA structure). Thus, the nucleotide sequence in one chain is complementary to, but not identical to, that in the other chain. The diameter of the double helix measured between phosphorus atoms is 2.0 nm. The pitch is 3.4 nm. There are 10 base pairs per turn. Thus the rise per base pair is 0.34 nm, and bases are stacked in the center of the helix. This form (B form), whose base pairs lie almost normal to the helix axis, is stable under high humidity and is thought to approximate the conformation of most DNA in cells. However, the base pairs in another form (A form) of DNA, which likely occurs in complex with histone, are inclined to the helix axis by about 20° with 11 base pairs per turn. While DNA molecules may exist as straight rods, the two ends bacterial DNA are often covalently joined to form circular DNA molecules, which are frequently supercoiled. 

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