- Email: [email protected]
Bacteriocins
Bacteriocins R Lagos, Universidad de Chile, Santiago, Chile © 2013 Elsevier Inc. All rights reserved.
Glossary ABC exporter Also called type I exporter, corresponds to membrane proteins that participate in the export of metabolites, peptides, or proteins in a specific manner (dedicated exporter, energy driven by adenosine triphosphate (ATP) hydrolysis), with a characteristic domain called ABC (ATP-binding cassette). Pathogenicity island Part of a bacterial genome acquired by horizontal transfer encoding genes involved in pathogenesis. Quorum sensing Communication between bacteria given by molecules secreted by the cells that coordinate the
Introduction Bacteriocins are a large family of ribosomally synthesized pro teinaceous toxins produced by bacteria and Archaea that have antimicrobial activity against bacteria closely related to the producer strain. The term ‘bacteriocin’ was introduced to denote a toxic protein or peptide produced by any type of bacteria that is active on related bacteria but does not harm the producing cell. This expression was inspired by analogy with colicin, the first bacteriocin described that was produced by Escherichia coli. In this case, the suffix cin was added to the producing species, such as in the case of pyocins (from Pseudomonas pyocyanea, now P. aeruginosa) and cloacins (Enterobacter cloacae), to mention a few. The genus name has also been employed to name bacteriocins, for instance klebi cins (from Klebsiella), lactococcins (from Lactococcus), staphylococcins (from Staphylococcus), and many others. Although bacteriocins present toxic activities on bacteria, they should not be confused with ‘toxins’ (exotoxins), a term that is reserved for proteins produced by bacteria that have a toxic effect on an animal host through specific cytotoxic action on specialized cells (e.g., hemolysins and cytolysins).
expression of certain genes according to the bacterial population density. Salmochelin Enterochelin (cyclic trimer of N-(2,3 dihydroxybenzoyl)-L-serine) glycosylated with one or two glucose molecules; a siderophore produced by Salmonella sp. and other pathogenic strains. Siderophore Compounds synthesized and secreted by microorganisms that chelate iron with high affinity. The siderophore–iron complex is recognized by specific receptors for an efficient internalization and release of iron.
associated to the TonB or Tol systems, both used for important biological functions involved in the import of molecules (nat ural ligands) such as vitamin B12, iron bound to siderophores, and others. Receptor recognition is key for the antibacterial activity, and is the reason why the Gram-negative bacteriocins in general have a narrow host range: they are active only on strains having the specific receptors. This is in contrast with classic antibiotics, which are directed toward a broader spec trum of bacteria. The situation as regards Gram-positive bacteria is different: in this case, the outer membrane barrier does not exist, and the cell wall allows the passage of relatively large molecules; hence, absorption shows little specificity. However, there is a family of Gram-positive bacteriocins that needs specific interaction with receptor proteins from the cyto plasmic membrane, and consequently has a very narrow spectrum of activity, whereas there are cationic bacteriocins that present an initial interaction with anionic cell-surface polymers (such as teichoic acid, lipoteichoic acid, and anionic phospholipids), which results in a class of bacteriocins that are active against a broad variety of bacteria belonging to different genera.
Immunity Mechanisms of Action The mechanisms by which bacteriocins exert antimicrobial activity are diverse. The most common are the disruption of the cytoplasmic membrane by pore-forming structures, cellwall interference by inhibition of peptidoglycan synthesis, nucleases production (DNases or RNases), nucleic acid metabolism interference by inhibition of gyrase or RNA poly merase, and others. In all these cases, the bacteriocin has to reach the target in the cell. In the case of Gram-negative bac teria, the target can be located at the periplasm, the cytoplamic membrane, or at the cytoplasm, and it is reached after a receptor-recognition step. The main two uptakes utilized by Gram-negative bacteriocins are outer membrane proteins
Brenner’s Encyclopedia of Genetics, 2nd edition, Volume 1
How does the producing strain avoid killing itself? There is a self-protection mechanism that prevents the toxic effect of the bacteriocin on the producing strain, a process called immunity (although it is very different from phage immunity). This protec tion is achieved by specific mechanisms that block the action of the bacteriocins. For example, the immunity for pore-forming bacteriocins is given by membrane proteins that neutralize the bacteriocin by a direct interaction, or by an interaction with proteins needed for the insertion of the bacteriocin into the membrane; the immunity to bacteriocins with nuclease activity (or other activities) is given by association of the immunity protein with the bacteriocin, forming an inactive complex. Immunity should not be confused with resistance. Although,
doi:10.1016/B978-0-12-374984-0.00291-6
277
278
Bacteriocins
in both cases, there is no toxic effect after the exposure to the bacteriocin, the immunity corresponds to a specific mechanism to counteract the toxic effect, whereas the resistance results, for example, because of the lack of a specific receptor on the target cell, so the bacteriocin cannot reach the objective.
Bacteriocin Produced by Gram-Negative Bacteria Two main groups can be distinguished: colicins or colicinslike bacteriocins, which are heat labile and have a high mole cular weight (>20 000); and small (<10 000), heat-resistant bacteriocins, which due to their small size are called microcins. Colicin and colicin-like bacteriocins are typically encoded in plasmids and the genetic organization is very simple, with only three components: the colicin, the immunity, and the lysis genes. The production of colicins occurs under stress conditions: they are induced by the SOS system, in which not only the expression of the colicin is induced but also the lysis gene, which in turn causes cellular lysis, with the libera tion of the bacteriocin. Hence, the release of colicins to the extracellular space implies bacterial death. Another salient feature of colicins is that they have three domains that are related with the steps necessary for antibacterial activity: reception (central domain), translocation (N terminal), and toxicity (C terminal). The first is needed for binding to the receptor at the outer membrane; the second for translocation across the cell envelope; and the third for the killing. In this process, colicins would unfold, and maintain an extended conformation across the cell envelope in contact with the receptor. Typical pore-forming colicins are colicins E1, A, B, K, Ia, Ib, and N; those with DNase activity are E2, E7, and E9; those with RNase activity (transfer RNA (tRNA) and 16S RNA) are D, E5, E3, E4, and E6; and that blocking peptidoglycan synthesis is colicin M. It is worth mentioning that most of the plasmid vectors utilized today carry the replication origin of plasmid ColE1, which encodes for colicin E1. Microcins can be either plasmid or chromosomally encoded. They are subdivided into very small (mass <5000) microcins that are highly modified, the most representative microcins being B17 (inhibits DNA gyrase), J25 (inhibits RNA polymerase), and C7 (affects protein synthesis), and microcins with a mass between 5 and 10 000 that may be or not posttranslationally modified. Microcins E492 (pore forming), H47, and M are modified at the C-terminal with salmochelin-like molecules, whereas microcin L and colicin V belong to the non-modified group. Microcins are heat resistant, usually induced in the stationary phase, and, in contrast to colicins, they are exported to the extracellular space by specific ABC (ATP-binding cassette) exporters encoded in the microcin gene clusters; consequently, the liberation of microcins does not result in cellular death. In this regard, they are similar to some bacteriocins produced by Gram-positive bacteria, as well as in other characteristics such as thermostability, hydrophobi city, resistance to extreme pH, and resistance to some proteases. Microcins M and H47 can be chromosomally encoded into pathogenicity islands.
Bacteriocins Produced by Gram-Positive Bacteria There is a great diversity of bacteriocins produced by Grampositive microorganisms. Interestingly, the term ‘bacteriocin’ is commonly associated with those produced by Gram-positive bacteria, in contrast to those produced by Gram-negative bac teria, which are mainly identified as ‘colicins’ and ‘microcins’. The main difference between bacteriocins from Gram-positive and colicins from Gram-negative bacteria are: their production is not lethal because they are secreted through specific or sec-dependent export pathways; and some Gram-positive bac teria have evolved bacteriocin-specific regulation (such as quorum sensing), in contrast to Gram-negative bacteriocins that are under the host’s regulatory networks. Gram-positive bacteriocins have been classified into four groups: class I, posttranslationally modified bacteriocins that contain modified amino acids, such as lanthionine, methyllanthionine, and dehy drated amino acids, and that are collectively known as lantibiotics; class II, small (<10 000) heat-stable, non-modified bacteriocins; class III, large (>10 000) heat-labile bacteriocins; and class IV, complex bacteriocins, carrying lipid or carbohydrate moieties. Class I, II, and III Gram-positive bacteriocins are further subdivided according to other characteristics. The most studied are class I and class II, many of them being produced by lactic acid bacteria (LAB), also known as LAB bacteriocins. Lantibiotics (or class I) are bacteriocins toxic on a broad range of Gram-positive bacteria, encoded in complex genetic systems located either in plasmids or in the chromosome in which are present the export and maturation genes needed for the specific exporters and for posttranslational modification. Lantibiotic production is regulated by a strategy called ‘quorum sensing’, in which the concentration of the bacteriocin functions as a signal molecule that evaluates cellular density, and after certain point induces its own expression. Lantibiotics are synthe sized as precursor peptides, with an N-terminal leader peptide that is processed. Several lantibiotics are produced by food-grade bacteria, which made them suitable as food preservatives. The most representative member of this group is nisin, a pore-forming bacteriocin with a broad variety of targets, among them strains of Lactococcus, Streptococcus, Staphylococcus, Listeria, Mycobacterium, Bacillus, and Clostridium. Nisin would use Lipid II, a peptidoglycan precursor, as a docking molecule for binding to specific membranes. Other lantibiotics are epider min, gallidermin, cytolysin, and cynnamicin. Class II bacteriocins encompass hydrophobic thermostable small non-modified bacteriocins produced by Lactobacillus, Lactococcus, Pediococcus, and Enterococcus among other strains. They function either as a single peptide or may require two or more components for the toxic activity. The bacteriocin is synthesized as a precursor that is processed while being exported by ABC exporters. Pediocins, listeriocins, lactococcins, and enterocins belong to this group.
Bacteriocins Produced by Archaea The proper name for bacteriocins produced by Archaea is ‘archaeocins’. The best-characterized member are the halocins, produced by halobacteria. They are produced mainly in the stationary phase.
Bacteriocins
Ecological Significance of Bacteriocins Although it is evident that bacteriocins serve some functions in bacterial communities, many studies have turned out to be inconclusive or contradictory. However, new develop ments point to the role of bacteriocins as regulators of bacterial populations. The rock–paper–scissor model (pair interactions between resistant, sensitive, and producing strains) lends support to the hypothesis that bacteriocins may promote rather than eliminate bacterial diversity in the ecosystem.
279
case of microcin E492, which is modified at the C-terminal end with a salmochelin-like molecule that is recognized by side rophore–iron receptors, and allow bacteriocin translocation to the target cell. Eventually, this knowledge may be used in the design of new antibiotics that need to cross the outer-membrane barrier. Microcin E492 has also antitumor properties that can be exploited in cancer therapy.
See also: Col Factors; Plasmids; Quorum Sensing; SOS Repair.
Further Reading Applications Application of bacteriocins in food preservation has been suc cessful: currently, the use of the lantibiotic nisin has been approved by the Food and Drug Administration (FDA) as a food preservative. In this regard, LAB bacteriocins would be advantageous because they are produced by bacteria that have been in food for centuries without producing any harm. Pharmaceutical applications for epidermin and gallidermin are been explored for acne. In another line of research, bacter iocins may be used as structural models in the design of new antibiotics, specifically if the toxic part of the molecule has been identified. The strategies used by bacteriocins may also be used as a model, such as the case of the Trojan horse strategy, in which the host is deceived by molecules that resemble normal compounds required for cellular growth. This is the
Cascales E, Buchanan SK, Duché D, et al., (2007) Colicin biology. Microbiology and Molecular Biology Reviews 71: 158–229. Gillor O, Nigro LM, and Riley MA (2005) Genetically engineered bacteriocins and their potential as the next generation of antimicrobials. Current Pharmaceutical Design 11: 1067–1075. Riley MA and Chavan MA (eds.) (2007) Bacteriocins: Ecology and Evolution. Berlin; Heidelberg: Springer-Verlag. Riley MA and Gillor O (eds.) (2007) Research and Applications in Bacteriocins. Norfolk: Horizon Bioscience. Riley MA and Wertz JE (2002) Bacteriocins: Evolution, ecology, and application. Annual Review of Microbiology 56: 117–137.
Relevant Websites http://bagel2.molgenrug.nl – Bagel2: The bacteriocin mining tool. BAGEL2 is a web-based bacteriocin mining tool.
Glossary ABC exporter Also called type I exporter, corresponds to membrane proteins that participate in the export of metabolites, peptides, or proteins in a specific manner (dedicated exporter, energy driven by adenosine triphosphate (ATP) hydrolysis), with a characteristic domain called ABC (ATP-binding cassette). Pathogenicity island Part of a bacterial genome acquired by horizontal transfer encoding genes involved in pathogenesis. Quorum sensing Communication between bacteria given by molecules secreted by the cells that coordinate the
Introduction Bacteriocins are a large family of ribosomally synthesized pro teinaceous toxins produced by bacteria and Archaea that have antimicrobial activity against bacteria closely related to the producer strain. The term ‘bacteriocin’ was introduced to denote a toxic protein or peptide produced by any type of bacteria that is active on related bacteria but does not harm the producing cell. This expression was inspired by analogy with colicin, the first bacteriocin described that was produced by Escherichia coli. In this case, the suffix cin was added to the producing species, such as in the case of pyocins (from Pseudomonas pyocyanea, now P. aeruginosa) and cloacins (Enterobacter cloacae), to mention a few. The genus name has also been employed to name bacteriocins, for instance klebi cins (from Klebsiella), lactococcins (from Lactococcus), staphylococcins (from Staphylococcus), and many others. Although bacteriocins present toxic activities on bacteria, they should not be confused with ‘toxins’ (exotoxins), a term that is reserved for proteins produced by bacteria that have a toxic effect on an animal host through specific cytotoxic action on specialized cells (e.g., hemolysins and cytolysins).
expression of certain genes according to the bacterial population density. Salmochelin Enterochelin (cyclic trimer of N-(2,3 dihydroxybenzoyl)-L-serine) glycosylated with one or two glucose molecules; a siderophore produced by Salmonella sp. and other pathogenic strains. Siderophore Compounds synthesized and secreted by microorganisms that chelate iron with high affinity. The siderophore–iron complex is recognized by specific receptors for an efficient internalization and release of iron.
associated to the TonB or Tol systems, both used for important biological functions involved in the import of molecules (nat ural ligands) such as vitamin B12, iron bound to siderophores, and others. Receptor recognition is key for the antibacterial activity, and is the reason why the Gram-negative bacteriocins in general have a narrow host range: they are active only on strains having the specific receptors. This is in contrast with classic antibiotics, which are directed toward a broader spec trum of bacteria. The situation as regards Gram-positive bacteria is different: in this case, the outer membrane barrier does not exist, and the cell wall allows the passage of relatively large molecules; hence, absorption shows little specificity. However, there is a family of Gram-positive bacteriocins that needs specific interaction with receptor proteins from the cyto plasmic membrane, and consequently has a very narrow spectrum of activity, whereas there are cationic bacteriocins that present an initial interaction with anionic cell-surface polymers (such as teichoic acid, lipoteichoic acid, and anionic phospholipids), which results in a class of bacteriocins that are active against a broad variety of bacteria belonging to different genera.
Immunity Mechanisms of Action The mechanisms by which bacteriocins exert antimicrobial activity are diverse. The most common are the disruption of the cytoplasmic membrane by pore-forming structures, cellwall interference by inhibition of peptidoglycan synthesis, nucleases production (DNases or RNases), nucleic acid metabolism interference by inhibition of gyrase or RNA poly merase, and others. In all these cases, the bacteriocin has to reach the target in the cell. In the case of Gram-negative bac teria, the target can be located at the periplasm, the cytoplamic membrane, or at the cytoplasm, and it is reached after a receptor-recognition step. The main two uptakes utilized by Gram-negative bacteriocins are outer membrane proteins
Brenner’s Encyclopedia of Genetics, 2nd edition, Volume 1
How does the producing strain avoid killing itself? There is a self-protection mechanism that prevents the toxic effect of the bacteriocin on the producing strain, a process called immunity (although it is very different from phage immunity). This protec tion is achieved by specific mechanisms that block the action of the bacteriocins. For example, the immunity for pore-forming bacteriocins is given by membrane proteins that neutralize the bacteriocin by a direct interaction, or by an interaction with proteins needed for the insertion of the bacteriocin into the membrane; the immunity to bacteriocins with nuclease activity (or other activities) is given by association of the immunity protein with the bacteriocin, forming an inactive complex. Immunity should not be confused with resistance. Although,
doi:10.1016/B978-0-12-374984-0.00291-6
277
278
Bacteriocins
in both cases, there is no toxic effect after the exposure to the bacteriocin, the immunity corresponds to a specific mechanism to counteract the toxic effect, whereas the resistance results, for example, because of the lack of a specific receptor on the target cell, so the bacteriocin cannot reach the objective.
Bacteriocin Produced by Gram-Negative Bacteria Two main groups can be distinguished: colicins or colicinslike bacteriocins, which are heat labile and have a high mole cular weight (>20 000); and small (<10 000), heat-resistant bacteriocins, which due to their small size are called microcins. Colicin and colicin-like bacteriocins are typically encoded in plasmids and the genetic organization is very simple, with only three components: the colicin, the immunity, and the lysis genes. The production of colicins occurs under stress conditions: they are induced by the SOS system, in which not only the expression of the colicin is induced but also the lysis gene, which in turn causes cellular lysis, with the libera tion of the bacteriocin. Hence, the release of colicins to the extracellular space implies bacterial death. Another salient feature of colicins is that they have three domains that are related with the steps necessary for antibacterial activity: reception (central domain), translocation (N terminal), and toxicity (C terminal). The first is needed for binding to the receptor at the outer membrane; the second for translocation across the cell envelope; and the third for the killing. In this process, colicins would unfold, and maintain an extended conformation across the cell envelope in contact with the receptor. Typical pore-forming colicins are colicins E1, A, B, K, Ia, Ib, and N; those with DNase activity are E2, E7, and E9; those with RNase activity (transfer RNA (tRNA) and 16S RNA) are D, E5, E3, E4, and E6; and that blocking peptidoglycan synthesis is colicin M. It is worth mentioning that most of the plasmid vectors utilized today carry the replication origin of plasmid ColE1, which encodes for colicin E1. Microcins can be either plasmid or chromosomally encoded. They are subdivided into very small (mass <5000) microcins that are highly modified, the most representative microcins being B17 (inhibits DNA gyrase), J25 (inhibits RNA polymerase), and C7 (affects protein synthesis), and microcins with a mass between 5 and 10 000 that may be or not posttranslationally modified. Microcins E492 (pore forming), H47, and M are modified at the C-terminal with salmochelin-like molecules, whereas microcin L and colicin V belong to the non-modified group. Microcins are heat resistant, usually induced in the stationary phase, and, in contrast to colicins, they are exported to the extracellular space by specific ABC (ATP-binding cassette) exporters encoded in the microcin gene clusters; consequently, the liberation of microcins does not result in cellular death. In this regard, they are similar to some bacteriocins produced by Gram-positive bacteria, as well as in other characteristics such as thermostability, hydrophobi city, resistance to extreme pH, and resistance to some proteases. Microcins M and H47 can be chromosomally encoded into pathogenicity islands.
Bacteriocins Produced by Gram-Positive Bacteria There is a great diversity of bacteriocins produced by Grampositive microorganisms. Interestingly, the term ‘bacteriocin’ is commonly associated with those produced by Gram-positive bacteria, in contrast to those produced by Gram-negative bac teria, which are mainly identified as ‘colicins’ and ‘microcins’. The main difference between bacteriocins from Gram-positive and colicins from Gram-negative bacteria are: their production is not lethal because they are secreted through specific or sec-dependent export pathways; and some Gram-positive bac teria have evolved bacteriocin-specific regulation (such as quorum sensing), in contrast to Gram-negative bacteriocins that are under the host’s regulatory networks. Gram-positive bacteriocins have been classified into four groups: class I, posttranslationally modified bacteriocins that contain modified amino acids, such as lanthionine, methyllanthionine, and dehy drated amino acids, and that are collectively known as lantibiotics; class II, small (<10 000) heat-stable, non-modified bacteriocins; class III, large (>10 000) heat-labile bacteriocins; and class IV, complex bacteriocins, carrying lipid or carbohydrate moieties. Class I, II, and III Gram-positive bacteriocins are further subdivided according to other characteristics. The most studied are class I and class II, many of them being produced by lactic acid bacteria (LAB), also known as LAB bacteriocins. Lantibiotics (or class I) are bacteriocins toxic on a broad range of Gram-positive bacteria, encoded in complex genetic systems located either in plasmids or in the chromosome in which are present the export and maturation genes needed for the specific exporters and for posttranslational modification. Lantibiotic production is regulated by a strategy called ‘quorum sensing’, in which the concentration of the bacteriocin functions as a signal molecule that evaluates cellular density, and after certain point induces its own expression. Lantibiotics are synthe sized as precursor peptides, with an N-terminal leader peptide that is processed. Several lantibiotics are produced by food-grade bacteria, which made them suitable as food preservatives. The most representative member of this group is nisin, a pore-forming bacteriocin with a broad variety of targets, among them strains of Lactococcus, Streptococcus, Staphylococcus, Listeria, Mycobacterium, Bacillus, and Clostridium. Nisin would use Lipid II, a peptidoglycan precursor, as a docking molecule for binding to specific membranes. Other lantibiotics are epider min, gallidermin, cytolysin, and cynnamicin. Class II bacteriocins encompass hydrophobic thermostable small non-modified bacteriocins produced by Lactobacillus, Lactococcus, Pediococcus, and Enterococcus among other strains. They function either as a single peptide or may require two or more components for the toxic activity. The bacteriocin is synthesized as a precursor that is processed while being exported by ABC exporters. Pediocins, listeriocins, lactococcins, and enterocins belong to this group.
Bacteriocins Produced by Archaea The proper name for bacteriocins produced by Archaea is ‘archaeocins’. The best-characterized member are the halocins, produced by halobacteria. They are produced mainly in the stationary phase.
Bacteriocins
Ecological Significance of Bacteriocins Although it is evident that bacteriocins serve some functions in bacterial communities, many studies have turned out to be inconclusive or contradictory. However, new develop ments point to the role of bacteriocins as regulators of bacterial populations. The rock–paper–scissor model (pair interactions between resistant, sensitive, and producing strains) lends support to the hypothesis that bacteriocins may promote rather than eliminate bacterial diversity in the ecosystem.
279
case of microcin E492, which is modified at the C-terminal end with a salmochelin-like molecule that is recognized by side rophore–iron receptors, and allow bacteriocin translocation to the target cell. Eventually, this knowledge may be used in the design of new antibiotics that need to cross the outer-membrane barrier. Microcin E492 has also antitumor properties that can be exploited in cancer therapy.
See also: Col Factors; Plasmids; Quorum Sensing; SOS Repair.
Further Reading Applications Application of bacteriocins in food preservation has been suc cessful: currently, the use of the lantibiotic nisin has been approved by the Food and Drug Administration (FDA) as a food preservative. In this regard, LAB bacteriocins would be advantageous because they are produced by bacteria that have been in food for centuries without producing any harm. Pharmaceutical applications for epidermin and gallidermin are been explored for acne. In another line of research, bacter iocins may be used as structural models in the design of new antibiotics, specifically if the toxic part of the molecule has been identified. The strategies used by bacteriocins may also be used as a model, such as the case of the Trojan horse strategy, in which the host is deceived by molecules that resemble normal compounds required for cellular growth. This is the
Cascales E, Buchanan SK, Duché D, et al., (2007) Colicin biology. Microbiology and Molecular Biology Reviews 71: 158–229. Gillor O, Nigro LM, and Riley MA (2005) Genetically engineered bacteriocins and their potential as the next generation of antimicrobials. Current Pharmaceutical Design 11: 1067–1075. Riley MA and Chavan MA (eds.) (2007) Bacteriocins: Ecology and Evolution. Berlin; Heidelberg: Springer-Verlag. Riley MA and Gillor O (eds.) (2007) Research and Applications in Bacteriocins. Norfolk: Horizon Bioscience. Riley MA and Wertz JE (2002) Bacteriocins: Evolution, ecology, and application. Annual Review of Microbiology 56: 117–137.
Relevant Websites http://bagel2.molgenrug.nl – Bagel2: The bacteriocin mining tool. BAGEL2 is a web-based bacteriocin mining tool.