Isolation of cell-free bacterial inclusion bodies - Microbial Cell
Bacterial Cell
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Transcript of Bacterial Cell
• Cytoplasm
• The cytoplasm, or protoplasm, of bacterial cells is where the functions for cell growth, metabolism, and replication are carried out.
•It is a gel-like matrix composed of water(4/5 th), enzymes, nutrients, wastes, and gases and contains cell structures such as ribosomes, a chromosome, and plasmids.
• The cell envelope encases the cytoplasm and all its components. Unlike the eukaryotic (true) cells, bacteria do not have a membrane enclosed nucleus. The chromosome, a single, continuous strands of DNA, is localized, but not contained, in a region of the cell called the nucleoid.
• Nucleoid
The nucleoid is a region of cytoplasm where the chromosomal DNA is located. It is not a membrane bound nucleus, but simply an area of the cytoplasm where the strands of DNA are found.
Ribosomes:•Consists of RNA and protein and they are abundant in cytoplasm and often grouped in long chains called polyribosomes.
•The relative sizes of ribosomes and their subunits can determined their sedimentation rate by centrifugation or subject to a centrifugal force.•Bacterial ribosomes have sedimentation rate of 70S; their subunits have rates of 30S and 50S. Streptomycin binds 70S ribosome and stops protein synthesis but it can not bind 80S ribosome of eukaryotes and thereby eukaryotic cell remains unaffected.
•The unit used to measure sedimentation velocity is Svedberg, larger the S value faster will be the sedimentation velocity. Sedimentation velocity depends upon both shape and size and hence it is not a measure of mass.
• Plasmids
Plasmids, are small, extrachromosomal genetic structures carried by many strains of bacteria. Like the chromosome, plasmids are made of a circular piece of DNA.
Plasmids have been shown to be instrumental in the transmission of special properties, such as antibiotic drug resistance, resistance to heavy metals, and virulence factors necessary for infection of animal or plant hosts.
• Flagella - Flagella (singular, flagellum) are hairlike structures that provide a mean of locomotion for those bacteria who have them.
• The flagella beat in a propeller-like motion to help the bacterium move toward nutrients; away from toxic chemicals; or, in the case of the photosynthetic cyanobacteria; toward the light. The species of bacteria, which are motile, move due to presence of flagella.
•Genetically determined as to number and location, a species with one flagellum is called monotrichous; with two (one on each end), amphitrichous or bipolar; with a tuft at one end called lophotrichous; and with many flagella, peritrichous.
•Prokaryotes are known to exhibit a variety of types of tactic behavior, i.e., the ability to move (swim) in response to environmental stimuli.
•chemotaxis- a bacterium can sense the quality and quantity of certain chemicals in their environment and swim towards them (if they are useful nutrients) or away from them (if they are harmful substances).
• Positive or negative chemotaxis, phototaxis, or even magnetotaxis can determine the direction of movement which alternates between swimming and tumbling motion.
•Pili or fimbriae
• (singular, pilus), small hairlike projections emerging from the outside cell surface.
•These outgrowths assist the bacteria in attaching to other cells and surfaces, such as teeth, intestines, and rocks.
•Without pili, many disease-causing bacteria lose their ability to infect because they're unable to attach to host tissue. Gonorrhea is one of the widest spread of the reportable diseases due to attachment fimbriae.
•There are some pili with a different function: they are only found on cells which can undergo a process known as conjugation, a means of genetic transfer among bacteria. Such pili are call “sex or F pili”.
Cell wall - Peptidoglycan layer
Gram –Negative bacteria
HHööltje 1998ltje 1998
Peptidoglycan: PBP activitiesPeptidoglycan: PBP activities
endopeptidase
DD-carboxypeptidase
D-Ala
Peptidoglycan-Gram Positive bacteria
Lipopolysaccharide (endotoxin) in Gram–negative bacteria
Lane 1 markerLane 2 O-antigen positive E. coli strainLane 3 O-antigen negative E. coli strain
Backbone composed of alternating pyrophosphate units (POP), linked to glucosamine (G) and it is flanked outward by repeating units of polysaccharides (O-antigen) and inward by long chain fatty acids (Lipid A)
•60-90% peptidoglycan and removal of cell wall makes protoplast which bursts easily by osmotic shock.•Retention of crystal violet is directly proportional to cell wall thickness.•Due to lack of periplasmic space the digestive enzymes and toxin are not retainedand they are released into the environment.
•Only 10-20% of the cell wall is peptidoglycan remainder consists of various proteins, lipids and polysaccharides.•Toxins and digestives enzymes are retained in periplasmic space for action, but these enzymes or toxins do not harm the organism that produced them.•After digestion of cell wall they form spheroplast (containing CM, OM)
•Cell wall is thick, contains 60% lipids and very less peptidoglycan.•The lipids make acid-fast organisms impermeable to most other strains and protect them from acids and alkalis.•The organism grow slowly because the lipids impede entry of nutrients into cell and cells spend large amount of energy to synthesize lipids.•They can be stained by Gram staining method and stained as Gram positive.
Mycobacteria
Nuclear region or nucleoid:Nucleoid is centrally located, contains DNA, RNA and proteins associated with it. Most of the bacterium contains one circular chromosome (except V. cholerae, which has two). Manipulation of the microbial genome/ chromosomes is the most important tool in any biotechnology industry.
Internal membrane systems:Photosynthetic and cyanobacteria contain internal membrane system (chromometaphore), which has membranes derived from cell membrane (CM) and pigments (helps to capture lights).
Nitrifying bacteria, soil organisms those convert nitrogen compounds into forms usable by green plants, also have internal membranes. They house enzymes and those are used in deriving energy from oxidation of nitrogen compounds.
Inclusion bodies:
Bacteria can have within their cytoplasm a variety of small bodies collectively referred to as inclusion bodies. Some are called granules and other are called vesicles.
Granules: Densely compacted substances without a membrane covering. Each granule contains specific substances, such as glycogen (glucose polymer) and polyphosphate (phosphate polymer, supplies energy to metabolic processes). Sulfur bacteria contains reserve granules of sulfur.
Polyphosphate granules are called volutin or metachromic granules, because they display metachromasia (metachromic granules exhibit different intensities of color). These granules are depleted in starvation.
Vesicles: Certain bacteria have specialized membrane enclosed structures called vesicles (or vacuoles). Some aquatic photosynthetic bacteria and cyanobacteria have rigid gas-filled vacuoles and it helps in floating at a certain level.Some magnetotactic bacterium, eg. Aquaspirilliummagnetotacticum, stores Magnetitite (Ferric oxide).The presence of such magnetic inclusions enablesthese bacteria to responds to magnetic fields.
•There are certain bacterial cells which are called vegetative having low level of metabolizing nutrients. Vegetative cells of Bacillus and Clostridium, produce resting stages called endospores.•A bacterium produces a single endospore, which merely helps the survival not reproduction. (But the fungal spores are for survival as well as reproduction).•Sporulation or endospore formation is a mean by which some bacteria prepare for the possibilities for adversities in future.
•Endospores form within the cells contain very less water and are highly resistant to heat, drying, acids, bases, certain disinfectants and even radiation. •It consists of a core, surrounded by a cortex, a spore coat and in some cases delicately thin layer called exosporium.•The core has an outer core wall, a cell membrane, nuclear region and other cell components. They contain dipicolinic acid (Pyridine-2,6-dicarboxylic acid, a metal chelating agent) and large amount of calcium ions.•Endospores are capable of survive a few months to several thousands of years in adverse environment exerting heat, cold, radiation stress.•When conditions become more favorable the spores germinate or begin to develop into functional vegetative cells.
Endospore:
Polar monotrichousPseudomonas
Polar amphitrichousSpirillium Lophotrichous
Spirillum
PeritrichousSalmonella
Flagella:
SEM of peritrichous strain Proteous
Note: bacteria without flagella are called atrichous.
In Gram-negative Bacteria
•The diameter of a prokaryotic flagellium is about 1/10 th of that of eukaryotic.
•It is made up of protein subunits called flagellin.
•Each flagellium is attached to cell membrane with the help of proteins other than flagellin.
•The basal region has a hook like structure and a complex basal body.
•The basal body consists of a central rod or shaft surrounded by a set of rings.
•Gram negative bacteria has a pair of rings embedded in the cell membrane and another pair of rings associated with the peptidoglycan and lipopolysaccharide layers.
•Gram-positive bacteria has one ring embedded in the cell membrane and another in the cell wall.
In Gram-positive Bacteria
BACTERIAL MOVEMENT
•Bacterial movement is produced through the action of the flagella .
• Bacteria move toward attractive stimuli and away from harmful substances and waste products in the process known as chemotaxis.
•Monotrichous bacteria move forward in a simple response to chemotactic stimuli by the counterclockwise rotation of the flagellum. This forward movement is termed the "run".
•Negative chemotaxis causes clockwise rotation of the flagellum and results in a random tumbling motion.
•Peritrichous bacteria move in a similar fashion, even though the situation is somewhat complicated by a requirement for bundling of the flagella to produce coordinated action during counterclockwise rotation. The "tumble" in peritrichous bacteria is the result of bundle disruption during clockwise flagellar rotation.
•Both run and tumble are generally random movement; no one direction of movement is more likely than any other direction.
bacterial movement contd.
•Runs lasts for an average of 1.0 sec, during which the bacterium swims about 10-20 times the length of its body.
•Tumbles last about 0.1 sec and no forward progress can be achieved.
•“Cruising speed” of a bacteria is about 10 body length/second which is almost equal to the flying speed for us.
•Bacterial chemotaxis is controlled by a complex series of events beginning with the binding of an attractant molecule to a cell surface chemoreceptor.
• Chemoreceptors are often clustered at the ends of rod-shaped cells like E. coli. Chemoreceptors do not influence flagellar motion directly, but convey information through a phosphorylation cascade required for signaling.
•Information about the environment can be translated into motion within 200 milliseconds. A return to steady-state is assured by a coordinated feedback loop that quickly causes a reversion to original levels of protein phosphorylation in the absence of stimuli.
Pilli or fimbrae
• Conjugation pilli found in certain group of bacteria.
• Used for genetic material transfer.
• The transfer process is called conjugation.
• Antibiotic resistance transfer.
• Atatchment pilli, Help bacteria adhare to the surfaces, such as cell surfaces and interface of water and air.
• It helps in colonization and pathogenicity.
