General introduction and applications of bacteriocins

1. General Introduction and
Applications of Bacteriocins
Subject: Advances of Microbiology
MPhil Microbiology
Nahima
Submitted To
Dr. Syed Muhammad Ali

2. Contents
•Introduction
•Mode of action
•Classification
•Applications

3. INTRODUCTION
• There are many antibacterial substances produced by animals, plants,
insects, and bacteria, such as hydrogen peroxide, fatty acids, organic acids,
ethanol, antibiotics, and bacteriocins.
• Bacteriocins are bactericidal antibiotic like substances.
• In nature it is protein which produced by many bacteria.
• It reduces the growth of bacterial strains that are similar or closely
associated.
• It is actually a proteinaceous toxin that is created as a tiny molecule by
bacteria.
• The toxin is regarded to be an antibiotic of narrow spectrum and it is
produced by a wide array of bacteria.

4. • Bacteriocins were first discovered by André Gratia in 1925.
• He was involved in the process of searching for ways to kill bacteria, which
also resulted in the development of antibiotics and the discovery
of bacteriophage, all within a span of a few years.
• He called his first discovery a colicine because it killed E. coli.
• Name bacteriocin was proposed by Jacob (1952) .
• Bacteriocins are given specific names based on the bacterial species of
origin, for example :-
• Colicins - E. coli
• Pyocins - P.aeruginosa
• Diphthericins - C.diphtheriae

5. Definition
• Bacteriocins are a kind of ribosomal synthesized antimicrobial
peptides produced by bacteria, which can kill or inhibit bacterial
strains closely-related or non-related to produced bacteria, but will not
harm the bacteria themselves by specific immunity proteins.
• Antimicrobial peptides (AMPs) or proteins produced by bacteria are
categorized as bacteriocins.

6. Structure and Mode of action of
Bacteriocins
• The structure of this toxin usually resembles the shape of a protein or
peptide. It consists of amino acid chains of different size.
• Pore formation
• Nuclease activity
• Peptidoglycanase activity
• Due to the great variety of their chemical structures, bacteriocins affect
different essential functions of the living cell (transcription,
translation, replication, and cell wall biosynthesis).

7. Pore formation
• The proposed mode of action for bacteriocins is an initial binding to
the bacterial membrane by electrostatic forces between the negatively
charged membrane lipids and the positive charges of bacteriocins.
Subsequently, bacteriocin insertion in the lipid bilayer takes place, and
by insertion of the transmembrane. Thus, pores are formed through the
bacteria membrane which becomes permeabilized. the formation of
pores or channels in the inner-membrane cause leakage of cytoplasmic
compounds, destruct electrochemical gradient, ion loss, and cell death.

8. Nuclease activity
• Nuclease is an enzyme that is responsible for breaking the bonds
between nucleotides in nucleic acids. nucleotides, are the small
subunits that make up large nucleic acids, such as DNA and RNA.
These are DNase, 16S rRNase, and tRNase to non-specifically digest
DNA and RNA of bacteria.
• DNAase phosphodiester linkages in the DNA backbone, thus
degrading DNA.
• RNAase is also a type of nuclease that catalyzes the degradation of
RNA.

9. Peptidoglycanase activity
• These proteins can digest the peptidoglycan, leading to an inability to
synthesize peptidoglycan and bacterial death.

11. Classification
• BACTERIOCINS FROM GRAM-NEGATIVE BACTERIA
• Colicins
• Microcins
• BACTERIOCINS FROM GRAM-POSITIVE BACTERIA
• Class I (modified peptides)
• Class II (unmodified peptides), and
• Class III (large proteins, heat unstable)

12. Bacteriocins from gram-negative bacteria
Colicins
• 25–80kDa high molecular weight colicins protein
• Colicin gene cluster encodes, toxin protein, Immunity protein, lysis gene
• Colicin regonition receptors and translocator protein must be present in
target bacteria.
• According to translocation across the outer membrane, GROUPA and
GROUP B.
• GROUPA, Tol protein system,encoded on small peptides, lysis gene
• GROUP B, Ton protein system,encoded on large plasmids, without lysis
gene.
• bactericidal mechanisms: (1) Pore-forming type colicins (2) Nuclease type
colicins (3) Peptidoglycanase type colicins

13. Colicins Antibacterial activity Molecular weight (Da) Producing strain
GROUPA
A Pore forming 62989 Citrobacter freudii
E1 Pore forming 57279 Escherichia coli
E2 DNase 61561 Escherichia coli
E8 DNase ∼70000 Escherichia coli
E6 16S rRNase 58011 Escherichia coli
E5 tRNase 58245 Escherichia coli
Shigella sonnei
GROUP B
B Pore forming 54742 Escherichia coli
D tRNase 74683 Escherichia coli
M Peptidoglycanase 29453 Escherichia coli

14. Microcins
• Low molecular weight ribosomal synthesized hydrophobic antimicrobial
peptides (<10kDa)
• No lysis gene, secreted outside bacteria
• Enterobacteriaceae showing great tolerance to heat, extreme pH, and
proteases.
• The bactericidal mechanisms of microcins are diverse, including the pore-
forming type, the nuclease type, such as DNase and RNase functions, and
inhibitors of protein synthesis or DNA replication.
• Class I <5kDa,post translational modified
• Class II 5–10kDa, IIa and IIb (require 3 genes, linear peptides without
modification)

15. Bacteriocins from gram-positive bacteria
• These bacteriocins are low molecular weight antimicrobial peptides
with less than 60 amino acids.
• In Gram-positive bacteria, lactic acid bacteria (LAB) are the typical
bacteria producing a variety of bacteriocins of different sizes,
structures, physicochemical properties, and inhibitory spectrum.
• Class I peptides are post-translationally modified bacteriocins with
less than 28 amino acids small membrane active peptides (<5kDa),
linear (membrane disrupting) or globular peptides (cellular enzymes)

16. • Class II bacteriocins are circular peptide bacteriocins, Class II
bacteriocins are 30–60 amino acids (<10kDa), heat tolerance,
unmodified, 5 sub classes IIa-IIe.
• Class III bacteriocins are large molecular weight (>30 kDa), heat
unstable proteins. Class III can be further subdivided into two distinct
groups. Group A bacteriocins are the bacteriolytic enzymes which
killing the sensitive strains by lysis of the cell wall, such as Enterolisin
A. Group B bacteriocins are non-lytic proteins such as Caseicin

17. Bacteriocins Molecular weight(Da) Producing Strain
CLASS I
Nisin A 3352 Lactococcus lactis subsp.
lactic
Nisin U 3092 Streptococcus uberis
CLASS II
Pediocin PA-1 4629 Pediococcus acidilactici
Enterocin AS-48 7149 Enterococcus faecalis
CLASS III
Caseicin 80 ∼42000 Lactobacillus casei
Enterolisin A 34501 Enterococcus faecalis

18. Application Of Bacteriocins
• Treatment of pathogen associated diseases
• Natural preservatives in food industries.
• Cleaning up environmental hazards
• As anticancer agent
• Applications of Bacteriocins in Livestock Health

19. As food preservative
• Bacteriocins have attracted considerable interest for their use as safe food preservatives,
as they are easily digested by proteolytic enzymes of the mammalian gastrointestinal
tract. Also, most bacteriocin producers belong to lactic acid bacteria.
• Producing strains should be exhibit a broad spectrum of inhibition against bacterial
activity
• It is proven non-toxic to humans, have no associated health risks
• Does not alter the nutritional properties of the food product.
• Effective at low concentration.
• Active under refrigerated storage
• Present beneficial effects (e.g., improve safety, quality, and flavor of foods), display heat
and pH stability.

20. Application of bacteriocins for
biopreservation
• The application of bacteriocins for biopreservation of foods usually
includes the following methods, bacteriocins, either purified or
excreted by bacteriocin producing strains.
• Inoculation of food with the bacteriocin-producer strain.
• Addition of purified or semi-purified bacteriocin as food additive and
• Use of a product previously fermented with a bacteriocin-producing
strain as an ingredient in food processing.

21. • Bacteriocins have been used in the biopreservation of various foods,
either alone or in combination with other methods of preservation.
hemical additives (such as sodium lactate, potassium diacetate, and
others), heating, and high-pressure treatments.
• Many studies showed the potential of applying bacteriocins or
bacteriocin-producing strains into foods, such as meat, dairy products,
fish, alcoholic beverages, salads, and fermented vegetables.

22. Applications of bacteriocin-producing LAB
in dairy products
Bacteriocin-producing
strain
Application technique Product Features
Staphylococcus
equorum WS 2733
Cheese was inoculated
with bacteriocin-
producing strains
Soft cheese L. monocytogenesgrowth
was completely inhibited
at low contamination
Enterococcus faecalis A-
48-32
Inoculation with
bacteriocin-producing
strain
Skimmed milk and fresh
cheese
Reduced Staphylococcus
aureus, but efficacy was
lower in cheese.
Lactobacillus
sakei subsp. sakei 2a
Cheese was inoculated
with bacteriocin-
producing strain
Cheese spread L. monocytogenes was
reduced at 4°C and
decreased at 15°C, after
22–28 days

23. Other applications of Bacteriocins
• Some bacteriocins, such as subtilosin A from Bacillus subtilis, were reported as
having anti-viral.
• A variety of degradative enzymes that are produced by Pseudomonas and are
capable of cleaning up environmental hazards such as oil spills and toxic chemical
waste site.
• Bacteriocins are reported to inhibit important animal and plant pathogens,such as
Shiga toxin-producing E. coli (STEC), enterotoxigenic E. coli (ETEC),
methicillin-resistant Staphylococcus aureus (MRSA), Agrobacterium, and
Brenneria spp.
• Feeding bacteriocin-producing bacteria for reducing the gastrointestinal
colonization of livestock by foodborne pathogens.
• Nisin was also applied in the treatment of respiratory tract infections. Some study
reported the capacity of nisin to develop resistance in respiratory tract to prevent
growth of resistant Staphylococcus aureus or Streptococcus pneumonia.

24. • Bacteriocins are able to selectively act against cancer cells, most likely
due to the distinctive differences in the membranes.
• In cancer therapy, some researches indicate that bacteriocins show
activity against tumor cells. Considering that bacteriocins are naturally
and legally added in foods, bacteriocins may be suitable as a potential
anti-tumor drugs.
• Colicins could act as an anti-cancer drugof moderate potential.
Supplements of bacteriocin-producing probiotics may be another way
to prevent cancer occurrence.
• In a recentstudy, found that nisin had capabilities to prevent cancer cell
growth.

25. THANK YOU