Microbial Genetics Conjugation PDF

Conjugation Conjugation During plasmid conjugation, DNA strands separate and one moves from the donor bacterium to the recipient, serving as templates for replication. Self-transmissible plasmids can transfer themselves and assist mobilizable plasmids, providing an advantage without burdeni...

Conjugation Conjugation During plasmid conjugation, DNA strands separate and one moves from the donor bacterium to the recipient, serving as templates for replication. Self-transmissible plasmids can transfer themselves and assist mobilizable plasmids, providing an advantage without burdening the host. Any bacterium with a self-transmissible plasmid can donate DNA, using structures like the sex pilus in gram-negative bacteria. Classification of Conjugation F plasmid, key in bacterial genetics, contains tra genes and supports the conjugation process. It reveals a complex, evolved transfer and maintenance system. Systems on the F plasmid stabilize it through partitioning and postsegregational killing, ensuring plasmid maintenance in daughter cell Structure of F plasmid The F plasmid, key in bacterial genetics, contains tra genes and supports the conjugation process. It reveals a complex, evolved transfer and maintenance system. Mechanism of DNA Transfer during Conjugation in Gram-Negative Bacteria The F plasmid consists of tra genes, making up about a third of it, which are vital for conjugation and related processes. Components of tra genes: The tra genes are divided into Dtr (DNA transfer and conjugal replication) and Mpf (mating pair formation) components. Mpf component's role: The Mpf component holds donor and recipient cells together, forming a channel for DNA transfer and initiating plasmid DNA transfer. Pilus Pilus-Prominent, tube-like feature of the Mpf structure, composed of pilin proteins with a central channel. Direct transfer of pilus is debated Coupling Proteins: Coupling proteins in the Mpf system communicate with the Dtr component to initiate DNA transfer, providing specificity and efficiency in the transfer process. DNA Transfer Mechanism: The coupling protein, bound to the membrane channel, acts as a DNA translocator, ensuring efficient DNA transfer during conjugation. Pilus Mpf Component Pilus assembly Other components Dtr component prepares plasmid DNA for transfer, with various proteins, including the relaxase. The relaxase (TraI in the F plasmid) makes a single-strand break at the oriT sequence, initiating transfer, and recircularizes DNA in the recipient cell. OriT site is crucial for plasmid transfer initiation and DNA recircularization Primase Primases are necessary for RNA primer creation in chromosomal and plasmid DNA replication, though not always needed in the donor due to the free 3′ hydroxyl end. Donor-Produced Primase: In plasmids like RP4 and ColIb-P9, primase produced in the donor can prime DNA replication in the recipient cell. Promiscuity and Adaptation: Plasmids make their own RNA primers to transfer into a wider range of bacterial species, especially when host cell primase does not recognize plasmid sequences. Male Specific Phages infect cells with conjugation-associated pili, using the pilus as their adsorption site. Mutations in tra genes needed for pilus assembly prevent phage infection, helping identify tra genes crucial for pilus expression. Intermittent expression: Plasmids irregularly express pili to protect against phage infection and avoid host detection. Efficiency of Transfer Many plasmid transfer systems are highly efficient, with almost 100% transfer under optimal conditions, aiding methods like gene cloning and transposon mutagenesis.  Efficiency of Transfer Regulation of tra Genes: Plasmids transfer efficiently only shortly after entering cells; tra genes are usually repressed, with occasional relief in a few cells allowing plasmid transfer. Triparental Matings: This principle underlies triparental matings, ensuring widespread plasmid distribution among bacterial populations. Interspecies Transfer of Plasmids Promiscuous plasmids: Plasmids like R388, RP4, and pKM101 can transfer DNA between unrelated species, including gram-negative and gram-positive bacteria, cyanobacteria, and even plant and yeast cells. likely plays a significant role in evolution, explaining the similarity of genes with related functions across different organisms. Interspecies Transfer of Plasmids Antibiotic resistance: Promiscuous plasmids, often carrying antibiotic resistance genes, have become prevalent due to the indiscriminate use of antibiotics in medicine and agriculture. Conjugation and Type IV Protein Secretion Conjugation and type IV protein secretion are interrelated; the coupling protein acts like a DNA pump in protein translocation. Type IV Secretion Systems: Used for DNA and protein transfer between cytoplasm and environment, in bacterial pathogens, and for transferring effector molecules to eukaryotic cells. Specificity and Function: Processes are very specific, involving pili and membrane structures, as seen in the T-DNA transfer system of Agrobacterium tumefaciens. Conjugation and Type IV Protein Secretion Conjugation and Type IV Protein Secretion Homologs in T-DNA Transfer System: Some pathogenic bacteria have analogous type IV secretion systems, sharing similarities with T-DNA systems like those of Agrobacterium. CagA Toxin System- Helicobacter pylori Pertussis Toxin System: Bordetella pertussis Legionella pneumophila Virulence System: Legionella pneumophila  Mobilizable Plasmids Mobilizable Plasmids: These plasmids can't transfer themselves but can be transferred by a self- transmissible plasmid in the same cell. Naturally Occurring Plasmids: Minimal mobilizable plasmids with only the oriT sequence don't occur naturally; they typically encode their own Dtr systems, including relaxase and helicase. Mob Genes: The tra genes of the Dtr system in mobilizable plasmids are called mob genes, which help expand the range of self-transmissible plasmids for mobilization. PLASMID MOBILIZATION IN BIOTECHNOLOGY Efficient DNA Introduction: Mobilization helps introduce foreign DNA into bacteria, with mob sites in cloning vectors making transfer efficient. Size Advantage: Mobilizable plasmids are smaller than self- transmissible ones, needing fewer genes, allowing them to be introduced into various bacteria using larger promiscuous plasmids. Chromosome Transfer by Plasmid Chromosome Transfer: Plasmids can sometimes transfer chromosomal DNA during conjugation, crucial for bacterial genetics. Plasmids integrating into chromosomes form Hfr strains (high-frequency recombination), transferring chromosomes during conjugation. Transfer of Chromosomal DNA by Integrated Plasmids Chromosomal DNA transfer in Hfr strains begins with plasmid integration, with the plasmid expressing tra genes and transferring DNA starting at oriT. Chromosome Mobilization: Tra functions can mobilize chromosomes if a mobilizable plasmid integrates or the chromosome contains the plasmid's oriT, aiding gene mapping across bacterial genera. Transfer of Chromosomal DNA by Integrated Plasmids Prime Factors: Conjugal plasmids that integrate and excise, containing chromosomal genes, are called prime factors (e.g., F′ factor, R′ factor). Chromosomal Deletion: Plasmid excision leads to chromosomal gene deletion; essential genes on the prime factor keep the bacterium alive. Size and Stability: Prime factors can be large and unstable; smaller ones transfer more easily, making recipient cells new donors. Transfer Systems of Gram-Positive Bacteria The oriT sequences in gram-positive plasmids are often similar to those in gram-negative bacteria, involving rolling-circle DNA replication. Stabilization Systems: Both types of bacteria have plasmids with postsegregational killing systems and partitioning systems, stabilizing the plasmid population. Simple Mpf systems and Carry virulence determinants and antibiotic resistance genes, beneficial to the host. Plasmid-Attracting Pheromones Pheromone-Driven Mating: Enterococcus faecalis excretes small peptide pheromones stimulating tra gene expression in neighboring plasmids, inducing aggregation and mating. Pheromone Production: Genes encoding pheromones are on the Enterococcus chromosome, producing active pheromones through proteolysis during export. Plasmid-Attracting Pheromones Pheromone Sensing: Requires specific surface and cytoplasmic proteins; plasmids express proteins for specific pheromones, named after the plasmid they attract (e.g., cAD1 for pAD1). Integrating Conjugative Elements (ICE) Unlike plasmids, ICEs are integrated into chromosomes but can still transfer themselves or mobilize other elements, often forming DNA islands in bacterial chromosomes. Excision and Transfer: ICEs must be excised from the host DNA, transferred to another cell, and integrated into the recipient's DNA, similar to the process seen in Tn916. Integrating Conjugative Elements (ICE) Thank You Fully Editable Shapes  Fully Editable Icon Sets: A You can Resize without losing quality You can Change Fill Color & Line Color FREE PPT TEMPLATES www.allppt.com  Fully Editable Icon Sets: B You can Resize without losing quality You can Change Fill Color & Line Color FREE PPT TEMPLATES www.allppt.com  Fully Editable Icon Sets: C You can Resize without losing quality You can Change Fill Color & Line Color FREE PPT TEMPLATES www.allppt.com