Linear plasmid

pSLA2

G.Thirupugal

Introduction-pSLA2

Name : pSLA2

Original host : Streptomyces rochei

Host range : Narrow host range

Copy number : low copy number(1 or 2)

Size : 17Kb

Geographical location : Japan

Genome map of plasmid pSLA2

1.Telomeres are present in both the ends.

2.Two rep genes are located in internal region of DNA.

3.Two cis acting sites are located in the rep genes

4.cis1 located within the region of the rep1 gene.

5.cis2 located within the region of rep2 gene.

Rep1 and Rep2 functions

1.Rep1 and Rep2 proteins are essential for plasmid DNA

replication and are encoded by the same transcript.

2.Rep1 is a DNA binding protein,where binding is specific

to the pSLA2 replication origin.

BssHII BsaAI BamHI BstxI AgeI BbsI ApaI SphI

Rep1 Rep2pSLA2

3.Rep2 contains two highly conserved NTP-binding

motifs typical of ATPase domains of DNA helicases.

4.Rep2 was enhanced by the presence of single-stranded

DNA.

5.Site directed mutagenesis showed that both the Rep1 and

Rep2 proteins are essential for pSLA2 DNA replication.

Cis- acting loci for pSLA2

Bcl1 BamH Sph1 Asp718

cis1 cis2

Bgl II

pSLA217Kb

Two cis acting loci was identified by the plasmid pSLA2.

It is required for replication.

Cis1 a segment located within the 864bp protein coding

region of the rep1 gene.

Cis2 a 430bp DNA segment positioned downstream of the

rep2 protein coding region.

Cis1 contains the plasmid pSLA2 replication origin.

The sequence characteristics of cis1is four directed

repeat followed by several runs of CT or GA rich

sequences .

Cis2 act like partition locus of plasmid pSC101 and the

locus of staphylococcus plasmid pT181.

1.Linear plasmid pSLA2 have protein attached to

covalently to their 5’ DNA termini.

2.The linear plasmid replicates bi-directional towards the

telomeres from a site located near the center of the

molecule.

3.The replication of mode pSLA2 the ability of lacking

telomeres to replicate as circular molecules.

The replication of linear plasmid pSLA2

4.The replication of pSLA2 is initiated at a series of direct

repeats located within a gene specifying an essential DNA-

binding protein.

5.That this gene (rep1) is part of a bi-cistronic Rep1 operon

that can function in trans to promote plasmid replication from

the centrally located origins.

6.The second gene(rep2)in this operon encodes a protein that

has single stranded DNA dependent ATPase activity

resembles prokaryotic DNA helicases.

7.The origin of replication region of this plasmid replicon

thus has features in common with the origin of temperate

bacteriophages of E.coli, Salmonella typhimurium, and

Bacillus subtlest.

8.Suggesting that pSLA2 may occupy an evolutionary

intermediate position between plasmids and phages.

Replication-pSLA2

A

B

C

13 bp

13 bp

Significance of plasmid pSLA2

They produces polyketide antibiotics

a)Lankacidin

b)lankamycin

These antibiotics are used for treatment of swine dysentery

caused by Treponema hydrodysenteriae.

This plasmid also encodes biosynthesis of carotenoid.