BACTERIAL BACTERIAL CONJUGATIONCONJUGATION

INTRODUCTIONINTRODUCTION

Bacteria posses basic three mechanisms of lateral or horizontal gene Bacteria posses basic three mechanisms of lateral or horizontal gene transfer. They are,transfer. They are,

Transformation- transfer of gene from one bacterium to other Transformation- transfer of gene from one bacterium to other without cell to cell contact.without cell to cell contact.

Transduction– transfer of gene from one bacterium to other via Transduction– transfer of gene from one bacterium to other via agency of phages.agency of phages.

Conjugation- transfer of gene from one bacterium to other by cell Conjugation- transfer of gene from one bacterium to other by cell to cell contact by conjugal pore. to cell contact by conjugal pore.

Conjugation was first discovered in 1946 by two renowned Conjugation was first discovered in 1946 by two renowned scientists of the time scientists of the time Joshua Lederberg and Edward Tatum..

Bacterial conjugation is often incorrectly regarded as the bacterial equivalentequivalent of sexual reproduction or mating. Bacterial Bacterial conjugation is merely a transfer of gene from one bacterium to conjugation is merely a transfer of gene from one bacterium to other unlike the fusion of gametes in sexual reproduction in other unlike the fusion of gametes in sexual reproduction in higher organisms.higher organisms.

DISTINGUISHING CHARACTERISTICS OF DISTINGUISHING CHARACTERISTICS OF CONJUGATION CONJUGATION

DNA transfer requires cell-cell contact.DNA transfer requires cell-cell contact. DNA transfer occurs via a conjugal pore.DNA transfer occurs via a conjugal pore. DNA transfer occurs in one direction - from donor to DNA transfer occurs in one direction - from donor to

recipient not vice versa recipient not vice versa DNA transfer does not require protein synthesis in donor.DNA transfer does not require protein synthesis in donor. DNA transfer requires energy in donor cell - primarily ATP.DNA transfer requires energy in donor cell - primarily ATP.

The F+ strain

The F- strain

The mating bridge

Fertility plasmidFertility plasmid

The E. coli fertility plasmid are most extensively studied plasmid. The general features of this plasmid are,

1. Large circular plasmid (100 kb)2. Only 60% genes has been mapped.3. 32 kb is organized as a unit to transfer its genome to another

bacteria (transfer region or tra genes)4. Two methods of replication:a. oriV as free plasmid (one copy/ bacterial chromosome)b. uses bacterial chromosomal origin when integrated (oriC); oriV is

suppressed.

mechanismmechanism

Plasmid replication requires a "mating bridge" between the donor Plasmid replication requires a "mating bridge" between the donor and recipient cells.and recipient cells.

Before the mating bridge can form: Before the mating bridge can form:

- donor must recognize recipient cell - donor must recognize recipient cell

- donor must make contact with recipient cell - donor must make contact with recipient cell

The conjugative functions of the F plasmid are specified by The conjugative functions of the F plasmid are specified by a cluster of at least 25 a cluster of at least 25 tratransfer (tra) genes. They determine:nsfer (tra) genes. They determine:

expression of F pili.expression of F pili. synthesis and transfer of DNA during matingsynthesis and transfer of DNA during mating interference with the ability of Finterference with the ability of F++ bacteria to serve as recipients. bacteria to serve as recipients.

Mechanism contd…Mechanism contd…

Each FEach F++bacterium contains F pilibacterium contains F pili

Binding of F pili to specific outer membrane proteinBinding of F pili to specific outer membrane protein

Intercellular cytoplasmic bridge formation

Transfer of single strand from donor to recipient

Transferred strand is converted into double stranded circle in recipient.

The copy is retained in donor

The newly synthesized double stranded DNA is called exogenote and native DNA as endogenote.

Domain interacting with outer membrane protein

3D X ray crystallographic structure of protein pilin

General representation of mechanism of conjugation

The relaxase and the relaxosomeThe relaxase and the relaxosome

When conjugation is initiated, via a mating signal, a relaxase When conjugation is initiated, via a mating signal, a relaxase enzyme (an endonuclease) creates a nick in one plasmid DNA enzyme (an endonuclease) creates a nick in one plasmid DNA strand at the strand at the oriorigin of gin of ttransfer, or oriT. ransfer, or oriT.

The relaxase may work alone or in a complex of over a dozen The relaxase may work alone or in a complex of over a dozen proteins, known collectively as a relaxosome. proteins, known collectively as a relaxosome.

In the F-plasmid system, the relaxase enzyme is called TraI and In the F-plasmid system, the relaxase enzyme is called TraI and the relaxosome consists of TraI, TraD, TraY, TraM, and the the relaxosome consists of TraI, TraD, TraY, TraM, and the iintegrated ntegrated hhost ost ffactor, IHF. actor, IHF.

The transferred, or T-strand, is unwound from the duplex plasmid The transferred, or T-strand, is unwound from the duplex plasmid and transferred into the recipient bacterium in a 5'-terminus to 3'-and transferred into the recipient bacterium in a 5'-terminus to 3'-terminus direction. terminus direction.

The remaining strand is replicated, either independent of The remaining strand is replicated, either independent of conjugative action (vegetative replication, beginning at the oriV) conjugative action (vegetative replication, beginning at the oriV) or in concert with conjugation (conjugative replication similar to or in concert with conjugation (conjugative replication similar to the rolling circle replication of lambda phage). the rolling circle replication of lambda phage).

Mechanism illustratedMechanism illustrated

Helps creating the ssDNA conformation

Sterically prepare the DNA for other proteins to interact in specific spots.

Initiates F plasmid transfer

Conjugation protein assemblyConjugation protein assembly

Domains of protein traiDomains of protein trai

Formation of Hfr strainFormation of Hfr strain

There are two mechanisms of There are two mechanisms of integration integration

-homologous recombination-homologous recombination

-transposition-transposition

Formation of F’ plasmidFormation of F’ plasmid

1. Plasmids that “leave” the genome carrying chromosomal DNA are known as prime factors.

2. They “leave” the chromosome by homologous recombination, resulting in a deletion in the chromosome

AnimationsAnimations

Host rangeHost range

Bacteria (same or different species)Bacteria (same or different species) YeastYeast Plant cellsPlant cells Mammalian cells Mammalian cells

conclusionconclusion

The genetic information transferred is often beneficial to The genetic information transferred is often beneficial to the recipient cell. Benefits may include antibiotic resistance, the recipient cell. Benefits may include antibiotic resistance, other xenobiotic tolerance, or the ability to utilize a new other xenobiotic tolerance, or the ability to utilize a new metabolite. Such beneficial plasmids may be considered metabolite. Such beneficial plasmids may be considered bacterial endosymbionts. Some conjugative elements may also bacterial endosymbionts. Some conjugative elements may also be viewed as genetic parasites on the bacterium, and be viewed as genetic parasites on the bacterium, and conjugation as a mechanism was evolved by the mobile element conjugation as a mechanism was evolved by the mobile element to spread itself into new hosts. to spread itself into new hosts.