Bacterial genetics

Bacterial Genetics

Bacterial genetics refers to the study of genetic processes and mechanisms in bacteria. It includes the investigation of how genetic information is stored, replicated, and inherited in bacterial cells, as well as the exploration of genetic variation, gene expression, and the transfer of genetic material between bacteria.

Nucleic acids- The two main types of nucleic acids are deoxyribonucleic acid (DNA) and ribonucleic acid (RNA). DNA and RNA are nucleic acids that carry out cellular processes, especially the regulation and expression of genes. DNA is the genetic material found in all living organisms, ranging from single-celled bacteria to multicelled mammals. In prokaryotes, the DNA is not enclosed in a membranous envelope, but rather free-floating within the cytoplasm.

Nucleotides- DNA and RNA are made up of monomers known as nucleotides. The nucleotides combine with each other to form a polynucleotide: DNA or RNA. Each nucleotide is made up of three components:

1. a nitrogenous base

2. a pentose (five-carbon) sugar

3. a phosphate group

Prokaryotic DNA- Prokaryotic DNA refers to the genetic material found in prokaryotic cells, which include bacteria and archaea. Unlike eukaryotic cells, prokaryotic cells do not have a nucleus, and their DNA is not contained within a membrane-bound organelle.

STRUCTURE OF PROKARYOTIC DNA

Prokaryotic DNA is typically a single, circular molecule called a chromosome. Unlike eukaryotic cells, prokaryotes or bacterial cells don’t have a membrane-bound nucleus. Instead, their genetic material can be found in a region of the cytoplasm called the nucleoid. A bacterial cell typically has only a single, coiled, circular chromosome. However, there are a few bacteria that have more than one. Vibrio cholerae, the bacterium that causes cholera, has two circular chromosomes. Each chromosome contains a molecule of DNA that is supercoiled and compacted by nucleoid-associated proteins (NAPs). Bacterial cells may have only one chromosome, but that one chromosome is a very long DNA molecule that must be condensed to fit inside a tiny space.

Thus, Bacteria have the same type of nucleic acids as our cells, only the arrangement is different. Bacteria have one double-stranded DNA chromosome that holds their genetic material. Instead of linear chromosomes like we have, bacterial chromosomes are circular

However, some prokaryotes may also have additional circular pieces of DNA called plasmids. Actually, plasmids are small, circular, extrachromosomal DNA molecules that can carry extra characters to prokaryotes, such as bacteria, such as antibiotic resistance or the ability to produce toxins. Bacterial geneticists study plasmids to understand their replication, transfer between bacterial cells, and the role they play in bacterial adaptation and evolution.

The size of prokaryotic genomes can vary significantly, ranging from a few hundred thousand base pairs to several million base pairs.

Gene- It is a basic unit of heredity and a sequence of nucleotides in DNA or RNA that encodes the synthesis of a gene product, either RNA or protein. It is a unit of hereditary information that occupies a fixed position (locus) on a chromosome. A gene is the basic physical and functional unit of heredity. Genes are made up of DNA. Some genes act as instructions to make molecules called proteins. However, many genes do not code for proteins. Each chromosome contains many genes.

Genes can acquire mutations in their sequence, leading to different variants, known as alleles, in the population. These alleles encode slightly different versions of a protein, which cause different phenotypical traits.

FUNCTIONS OF PROKARYOTIC DNA

Prokaryotic DNA serves several important functions in prokaryotic cells, like

Genetic Information Storage: Prokaryotic DNA contains the complete set of genetic instructions necessary for the development, growth, and functioning of the prokaryotic cell. It carries the information required to synthesize all the proteins and functional RNA molecules needed by the cell.

Replication: Prokaryotic DNA undergoes replication to transfer genetic information to daughter cells during cell division. Replication involves the synthesis of a complementary strand of DNA for each existing strand, resulting in two identical copies of the DNA molecule.

Gene Expression: Prokaryotic DNA contains genes that are transcribed into RNA molecules, which are then translated into proteins. Gene expression is the process by which genetic information is used to synthesize functional molecules. Prokaryotic DNA serves as a template for transcription, where RNA molecules are produced from specific regions of the DNA sequence.

Regulation of Gene Expression: Prokaryotic DNA plays a crucial role in regulating the expression of genes. Bacteria have mechanisms to control when and to what extent genes are transcribed and translated. Regulatory regions on the DNA interact with regulatory proteins and other molecules to modulate gene expression in response to environmental signals and cellular needs.

Genetic Variation and Evolution: Prokaryotic DNA is subject to genetic variation, which contributes to the evolution of bacteria. Mutations, genetic rearrangements, and horizontal gene transfer events can introduce changes in the DNA sequence. This genetic variation can lead to the emergence of new traits, such as antibiotic resistance, virulence factors, or metabolic capabilities, allowing bacteria to adapt and survive in different environments.

Horizontal Gene Transfer: Prokaryotic DNA can be transferred horizontally between different bacteria, allowing the exchange of genetic material. Horizontal gene transfer can occur through processes like transformation (uptake of DNA from the environment), transduction (transfer of DNA by bacteriophages), and conjugation (direct transfer of DNA between cells). This mechanism plays a significant role in bacterial evolution, as it enables the spread of advantageous genes, such as antibiotic resistance genes, among bacterial populations.

Maintenance and Packaging: Prokaryotic DNA is organized and compacted within the nucleoid region of the cell. It associates with proteins that help maintain its structure and ensure proper DNA packaging. These proteins help organize the DNA molecule and prevent it from tangling or breaking.

GENETIC CODE

The genetic code may be defined as the exact sequence of DNA nucleotides read as three-letter words or codons that determines the sequence of amino acids in protein synthesis.

Properties of Genetic Code

The genetic code has some important properties.

1. The genetic code is a Triplet.

Singlet and doublet codes are not adequate to code for 20 amino acids; therefore, it was pointed out that the triplet code is the minimum required. The triplet code has 64 codons, which are sufficient to code for 20 amino acids and also for start and stop signals in the synthesis of a polypeptide chain. In a triplet code, three RNA bases code for one amino acid.

2.     The genetic code is Universal

The genetic code is nearly universal, meaning that the same codons encode the same amino acids across different organisms. From bacteria to plants to animals, the basic principles of the genetic code remain conserved.

3. The genetic code is comma-less

There is no signal to indicate the end of one codon and the beginning of the next. In other words, the codons are continuous and there are no demarcation lines between codons.

4. The genetic code is Non-overlapping.

A non-overlapping code means that the same letter is not used for two different codons. In other words, no single base can take part in the formation of more than one codon. The adjacent codons do not overlap. Example: There are Bases: CATGAT
Non-overlapping Code: 2, that is, CAT and GAT;
Overlapping Code: 4 that is CAT, GAT, ATG, and TAT

5. The genetic code is Non-ambiguous.

The genetic code has 64 codons. A particular codon will always code for the same amino acid. Out of these, 61 codons code for 20 different amino acids. However, none of the codons codes for more than one amino acid. In other words, each codon codes only for one amino acid. The same codon shall not code for two or more different amino acids (non-ambiguous). In case of ambiguous code, one codon should code for more than one amino acid. In the genetic code there is no ambiguity.

6. The genetic code is Redundant/degenerate.

The genetic code is degenerate, meaning that multiple codons can code for the same amino acid. Most amino acids are specified by more than one codon, except for methionine and tryptophan, which have a single codon each. Nine amino acids are coded by two codons each, one amino acid [Isoleucine] by three codons, five amino acids by 4 codons each, and three amino acids by 6 codons each. This multiple system of coding is known as a degenerate or redundant code system. Such a system provides protection to the organism against many harmful mutations, because if one base of a codon is mutated, there are other codons which will code for the same amino acid, and there will be no alteration in the polypeptide chain.

7. The genetic code has polarity.

The genetic code has polarity, that is, the code is always read in a fixed direction, i.e., in the 5′ → 3′ direction. It is apparent that if the code is read in the opposite direction (i.e., 3′ → 5′), it would specify 2 different proteins, since the codon would have a reversed base sequence. This is well known that the message in mRNA is read in the 5 -3 direction. Thus, the polarity of the genetic code is from 5' end to 3' end.

Plasmid

A plasmid is an extrachromosomal genetic element found in bacteria. Plasmids are small, circular DNA molecules that exist independently of the chromosomal DNA. They can replicate autonomously and can be transferred between different bacterial cells.




There are several types of plasmids based on their characteristics and functions.

• Fertility (F) Plasmids: F-plasmids, also known as conjugative plasmids, carry genes responsible for conjugation, a process by which genetic material is transferred between bacterial cells. These plasmids facilitate the transfer of genetic material, including the plasmid itself, from donor cells (F+) to recipient cells (F-).

• Resistance (R) Plasmids: Resistance plasmids contain genes that provide resistance to antibiotics or other toxic substances. These plasmids carry genes encoding enzymes, efflux pumps, or other mechanisms that enable bacteria to survive in the presence of antibiotics. Resistance plasmids contribute to the spread of antibiotic resistance among bacterial populations.

• Col Plasmids: Col plasmids produce colicins, which are bacteriocins that can kill or inhibit the growth of closely related bacterial strains. These plasmids enhance the survival of the host bacterium by producing antimicrobial substances that give them a competitive advantage over other bacteria.

• Virulence Plasmids: Virulence plasmids carry genes that contribute to the pathogenicity of bacteria. They encode factors such as toxins, adhesins, or proteins that help the bacteria evade the host immune system or facilitate colonization and infection. Virulence plasmids are often found in pathogenic bacteria and play a crucial role in causing disease.

• Degradative Plasmids: Degradative plasmids contain genes encoding enzymes that allow bacteria to break down and utilize complex substances such as hydrocarbons, pesticides, or toxic compounds. These plasmids enable bacteria to degrade and survive in environments polluted with specific substances that would otherwise be toxic to them.

• Cryptic Plasmids: Cryptic plasmids are small plasmids that do not confer any known phenotypic traits to the host bacterium. Their function and significance are often unclear, and they are referred to as "cryptic" because their role remains undiscovered.

Bacterial Recombination

Bacterial recombination is a type of genetic recombination in bacteria characterized by DNA transfer from one organism, called donor, to another organism, as the recipient. Horizontal gene transfer, also known as lateral gene transfer, is a process in which an organism transfers genetic material to another organism that is not its offspring. So simply, Genetic recombination is the transfer of DNA from one organism (donor) to another recipient. The transferred donor DNA may then be integrated into the recipient's nucleoid by various mechanisms (homologous, non-homologous). Genetic recombination of bacteria includes

a. Transformation
b. Transduction
c. Conjugation

Transformation
Transformation involves the uptake of free or naked DNA released by the donor by a recipient. Bacterial transformation is a process of horizontal gene transfer by which some bacteria take up foreign genetic material (naked DNA) from the environment. It was first reported in Streptococcus pneumoniae by Griffith in 1928.

The process of gene transfer by transformation does not require a living donor cell, but only requires the presence of persistent DNA in the environment. The prerequisite for bacteria to undergo transformation is their ability to take up free, extracellular genetic material. Such bacteria are termed competent cells.

The factors that regulate natural competence vary between various genera. Once the transforming factor (DNA) enters the cytoplasm, it may be degraded by nucleases if it is different from the bacterial DNA. If the exogenous genetic material is similar to bacterial DNA, it may integrate into the chromosome. Sometimes, the exogenous genetic material may co-exist as a plasmid with chromosomal DNA.

It was the first example of genetic exchange in bacteria to have been discovered. This was first demonstrated in an experiment conducted by Griffith in 1928. The presence of a capsule around some strains of pneumococci gives the colonies a glistening, smooth (S) appearance, while pneumococci lacking capsules produce rough (R) colonies. Strains of pneumococci with a capsule (type I) are virulent and can kill a mouse, whereas strains lacking it (type II) are harmless. Griffith found that mice died when they were injected with a mixture of live non-capsulated (R, type II) strains and heat-killed capsulated (S, type I) strains. Neither of these two, when injected alone, could kill the mice; only the mixture of the two proved fatal. Live S strains with capsule were isolated from the blood of the animal, suggesting that some factor from the dead S cells converted the R strains into S type.

Transformation: Genetic recombination in which a DNA fragment from a dead, degraded bacterium enters a competent recipient bacterium and it is exchanged for a piece of the recipient's DNA. It involves 4 steps.











Mechanism of transformation
Induction of Competence: Bacteria must first be made "competent" to take up DNA. This can occur naturally in some bacteria under certain conditions (natural competence) or can be induced in the laboratory by treatment with specific chemicals, electric fields, or through genetic manipulation (artificial competence).

 
DNA Uptake: Competent bacteria are then exposed to DNA from an external source, such as DNA released by lysed bacterial cells or purified plasmid DNA introduced into the environment.

 
Binding and Entry: The foreign DNA binds to receptors on the surface of the competent bacteria and is taken up into the bacterial cell. This process is facilitated by proteins (DNA-binding proteins) and structures on the bacterial cell surface.

 
RecA-Mediated Recombination: Once inside the bacterial cell, the foreign DNA may integrate into the bacterial chromosome through a process called homologous recombination. RecA, along with other proteins involved in recombination and repair, helps to facilitate this integration.

 
Expression of New Genes: If the integrated DNA contains functional genes, these genes can be expressed by the bacterial cell. This can lead to the acquisition of new traits or phenotypes conferred by the foreign DNA.


Transduction

Transduction is a process of genetic recombination in bacteria in which genes from a host cell (a bacterium) are incorporated into the genome of a bacterial virus (bacteriophage) and then carried to another host cell when the bacteriophage initiates another cycle of infection. In general transduction, any of the genes of the host cell may be involved in the process; in special transduction, however, only a few specific genes are transduced.

Genetic recombination in which a DNA fragment is transferred from one bacterium to another by a bacteriophage

• There are two types of transduction:

– Generalized transduction: A DNA fragment is transferred from one bacterium to another by a lytic bacteriophage that is now carrying donor bacterial DNA due to an error in maturation during the lytic life cycle.

– Specialized transduction: A DNA fragment is transferred from one bacterium to another by a temperate bacteriophage that is now carrying donor bacterial DNA due to an error in spontaneous induction during the lysogenic life cycle

Conjugation

Conjugation is the transfer of DNA from a donor to a recipient by direct physical contact between the cells. (Involves cell-to-cell contact). Bacterial conjugation is the transfer of genetic material between bacterial cells by direct cell-to-cell contact or by a bridge-like connection between two cells. This takes place through a pilus. In bacteria, there are two mating types: a donor (male) and a recipient (female), and the direction of transfer of genetic material is one way; DNA is transferred from a donor to a recipient.

F+ Cells:

These are the bacterial cells that contain the F plasmid. They are designated F+ simply because they have an F plasmid. We know a plasmid is an extrachromosomal DNA that can replicate independently. It is called the F plasmid because it has the F factor, which is the Fertility factor. This fertility factor contains the genes required for the transfer or conjugation.

F+ Cells = Cells containing F plasmid (F plasmid = Plasmid containing F factor)

F- Cells:

F- cells are the cells without the F plasmid. These cells act as recipient cells because they don’t have the F plasmid, and thus, they cannot donate the genetic material. They are designated as F- simply because they do not have the F plasmid.

F- Cells = Cells lacking the F plasmid

The process was discovered by Joshua Lederberg and Edward Tatum in 1946.

General mechanism of Conjugation

1. Donor cell produces a pilus.

1. Pilus attaches to the recipient cell and brings the two cells together.

2. The mobile plasmid is nicked, and a single strand of DNA is then transferred to the recipient cell.

3. Both cells synthesize a complementary strand to produce a double-stranded circular plasmid and also reproduce pili; both cells are now viable donors for the F-factor.

Mechanism of conjugation

a. F+ X F- crosses

i) Pair formation: The tip of the sex pilus comes in contact with the recipient, and a conjugation bridge is formed between the two cells. It is through this bridge that the DNA will pass from the donor to the recipient. Thus, the DNA is protected from environmental nucleases. The mating pairs can be separated by shear forces, and conjugation can be interrupted. Consequently, the mating pairs remain associated for only a short time.

ii) DNA transfer: The plasmid DNA is nicked at a specific site called the origin of transfer and is replicated by a rolling circle mechanism. A single strand of DNA passes through the conjugation bridge and enters the recipient, where the second strand is replicated.

iii) Result: This process explains the characteristics of F+ X F- crosses. The recipient becomes F+, the donor remains F+, and there is a low frequency of transfer of donor chromosomal genes. Indeed, as depicted in Figure & there is no transfer of donor chromosomal genes. In practice, however, there is a low level of transfer of donor chromosomal genes in such crosses.

b. Hfr X F- crosses

i) Pair Formation: Same as in F+ X F- crosses

ii) DNA transfer: The DNA is nicked at the origin of transfer and is replicated by a rolling circle mechanism. But the DNA that is transferred first is the chromosome. Depending upon where in the chromosome the F factor has integrated and in what orientation, different chromosomal genes will be transferred at different times. However, the relative order and distances of the genes will always remain the same. Only when the entire chromosome is transferred will the F factor be transferred. Since shearing forces separate the mating pairs it is rare that the entire chromosome will be transferred. Thus, the recipient does not receive the F factor in a Hfr X F- cross.

iii) Result: This mechanism explains the characteristics of Hfr X F- crosses. The recipient remains F-, the donor remains Hfr and there is a high frequency of transfer of donor chromosomal genes.

c.

F' X F- crosses

i) Pair formation: Same as in F+ X F- crosses

ii) DNA transfer: This process is similar to F⁺ X F^- crosses. However, since the F' has some chromosomal genes on it, these will also be transferred.

iii) Result: Homologous recombination is not necessary, although it may occur. This mechanism explains the characteristics of F' X F- crosses. The F- becomes F', the F' remains F', and there is a high frequency transfer of donor genes on the F' but a low frequency transfer of other donor chromosomal genes.