Halocins: salt-dependent bacteriocins produced by extremely halophilic rods

Rodrı́guez-Valera, Francisco; Juez, G.; Kushner, D. J.

doi:10.1139/m82-019

Cited by 94 publications

(82 citation statements)

References 7 publications

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“…As such, halocins are the haloarchaeal equivalent of the well-characterized protein antibiotics called bacteriocins that are produced by some 30 genera of the domain Bacteria (2). The term "halocin" was coined in 1982 by Francisco Rodriguez-Valera, who assumed that these substances were bacteriocins and who logically applied a similar nomenclature (28). Although archaeons share a "prokaryotic" cellular organization and morphological motif with members of the domain Bacteria, they are in separate domains and are only distantly related (41).…”

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Halocin S8: a 36-Amino-Acid Microhalocin from the Haloarchaeal Strain S8a

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2000

J Bacteriol

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Halocin S8 is a hydrophobic microhalocin of 36 amino acids (3,580 Da) and is the first microhalocin to be described. This peptide antibiotic is unique since it is processed from inside a much larger, 33,962-Da pro-protein. Halocin S8 is quite robust, as it can be desalted, boiled, subjected to organic solvents, and stored at 4°C for extended periods without losing activity. The complete amino acid sequence of halocin S8 was obtained first by Edman degradation of the purified protein and verified from the halS8 gene: H 2 N-S-D-C-N-I-N-S-N-T-A-A-D-V-I-L-C-F-N-Q-V-G-S-C-A-L-C-S-P-T-L-V-G-G-P-V-P-COOH.The halS8 gene is encoded on an ϳ200-kbp megaplasmid and contains a 933-bp open reading frame, of which 108 bp are occupied by halocin S8. Both the halS8 promoter and the "leaderless" halS8 transcript are typically haloarchaeal. Northern blot analysis revealed three halS8 transcripts: two abundant and one minor. Inspection of the 3 end of the gene showed only a single, weak termination site (5-TTTAT-3), suggesting that some processing of the larger transcripts may be involved. Expression of the halS8 gene is growth stage dependent: basal halS8 transcript levels are present in low concentrations during exponential growth but increase ninefold during the transition to stationary phase. Initially, halocin activity parallels halS8 transcript levels very closely. However, when halocin activity plateaus, transcripts remain abundant, suggesting inhibition of translation at this point. Once the culture enters stationary phase, transcripts rapidly return to basal levels.

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Halocin S8: a 36-Amino-Acid Microhalocin from the Haloarchaeal Strain S8a

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2000

J Bacteriol

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“…Halocins are haloarchaeal equivalents of eubacterial bacteriocins (2) and were first discovered in 1982 by F. RodriguezValera (24). Although nearly universal in halobacterial rods (17,32), only three halocins have been characterized in any detail: halocin H4 from Haloferax mediterranei R4 (15,16), halocin H6 from Haloferax gibbonsii Ma2.39 (30,31), and halocin HalR1 from Halobacterium sp.…”

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Isolation, sequence, and expression of the gene encoding halocin H4, a bacteriocin from the halophilic archaeon Haloferax mediterranei R4

et al. 1997

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The first gene to encode a haloarchaeal bacteriocin (halocin H4) has been cloned and sequenced from Haloferax mediterranei R4. Both the signal sequence in the halocin H4 preprotein and the monocistronic halH4 gene have some unusual features. The physiology of halH4 expression reveals that although halH4 transcripts are present at low basal levels during exponential growth, halocin H4 activity first appears as the culture enters stationary phase. As halocin activity levels increase, so do transcript levels, but then activity levels decrease precipitously while transcript levels remain elevated.Halocins are haloarchaeal equivalents of eubacterial bacteriocins (2) and were first discovered in 1982 by F. RodriguezValera (24). Although nearly universal in halobacterial rods (17, 32), only three halocins have been characterized in any detail: halocin H4 from Haloferax mediterranei R4 (15, 16), halocin H6 from Haloferax gibbonsii Ma2.39 (30, 31), and halocin HalR1 from Halobacterium sp. strain GN101 (23). In parallel with antibiotic production in the domain Bacteria (33), halocins H4, S8, and HalR1 are all initially detected as the cultures leave exponential growth and enter stationary phase (21). Consequently, we chose halocins, beginning with halocin H4, as models to study stationary-phase gene expression in the haloarchaea.Isolation of the halH4 gene. Halocin H4 was purified from H. mediterranei R4 (ATCC 33500) culture supernatants essentially as described elsewhere (4, 15). The amino-terminal sequences of the secreted protein and of two tryptic fragments (numbers 9 and 17 [ Fig. 1]) were used to design inosinecontaining degenerate oligodeoxynucleotide primers. Using these primers, we amplified the 5Ј end of the halocin H4 gene (halH4) by PCR and used the larger product (150 bp) as a probe to recover the gene from an enriched HindIII plasmid library.The start site of transcription of the halH4 gene was determined by primer extension (26). Note that the initiator AUG codon is only 4 bases from the 5Ј end of the message (Fig. 1). Similar leaderless transcripts are produced by other haloarchaeal genes, including bop (7); brp (3); hop (5); and arcA, arcB, and arcC (25). Inspection of the DNA sequence upstream of the halH4 transcriptional start site reveals a box A haloarchaeal promoter hexamer (Fig.

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“…Further examination of the cytocidal TuA4 supernatant revealed that the lethal component was a secreted protein, since haloarchaeal cell lysates were not toxic and the toxic activity could be abolished by thermophilic protease treatment. Peptide antibiotics have been identified as excreted products for numerous haloarchaea (29) and have been termed halocins (22) or microhalocins (20). The mechanism of action and target are known only for halocin H6, a Na ϩ /H ϩ antiporter inhibitor (16).…”

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“…For example, Sulfolobus islandicus produces an insoluble cell-associated peptide, termed a sulfolobicin, which is specific only for closely related species (19). Halophilic archaea secrete peptide antibiotics called halocins (or, if small, microhalocins) upon entry into stationary phase and in some cases after entry into stationary phase (20,22,26,28,29).…”

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Secreted Euryarchaeal Microhalocins Kill Hyperthermophilic Crenarchaea

et al. 2001

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Few antibiotics targeting members of the archaeal domain are currently available for genetic studies. Since bacterial antibiotics are frequently directed against competing and related organisms, archaea by analogy might produce effective antiarchaeal antibiotics. Peptide antibiotic (halocin) preparations from euryarchaeal halophilic strains S8a, GN101, and TuA4 were found to be toxic for members of the hyperthermophilic crenarchaeal genus Sulfolobus. No toxicity was evident against representative bacteria or eukarya. Halocin S8 (strain S8a) and halocin R1 (strain GN101) preparations were cytostatic, while halocin A4 (strain TuA4) preparations were cytocidal. Subsequent studies focused on the use of halocin A4 preparations and Sulfolobus solfataricus. Strain TuA4 cell lysates were not toxic for S. solfataricus, and protease (but not nuclease) treatment of the halocin A4 preparation inactivated toxicity, indicating that the A4 toxic factor must be a secreted protein. Potassium chloride supplementation of the Sulfolobus assay medium potentiated toxicity, implicating use of a salt-dependent mechanism. The utility of halocin A4 preparations for genetic manipulation of S. solfataricus was assessed through the isolation of UV-induced resistant mutants. The mutants exhibited stable phenotypes and were placed into distinct classes based on their levels of resistance.Small subunit (16S) rRNA sequence comparisons have identified a unique lineage or domain of prokaryotic organisms called archaea, which are currently subdivided into the euryarchaea, crenarchaea, and korarchaeota (1,33,34). Although prokaryotic in morphology, archaea employ eukaryotic mechanisms for many subcellular processes (32), including replication (2), transcription (12,14,15), and translation (5, 13). Cultivated archaea are further divided into the prominent biotypes of methanogens, halophiles (haloarchaea), and hyperthermophiles. Haloarchaea are members of the euryarchaea and thrive under conditions of high salt, while hyperthermophiles can be found in both euryarchaeal and crenarchaeal branches of the archaeal domain. Sulfolobus solfataricus is a hyperthermophilic aerobic crenarchaeote that inhabits acidic terrestrial hot springs. It is capable of both lithoautotrophic growth through sulfur oxidation (27, 35) and heterotrophic growth using a variety of defined carbon and energy sources (6,8,9,23).Antibiotics are broadly defined as natural, semisynthetic, and wholly synthetic substances that kill or inhibit the growth of microorganisms at low concentrations. One abundant class of bacterial antibiotics is the bacteriocins (3,7,11,25). These secreted proteinaceous compounds are produced by both gram-positive and gram-negative bacteria, range in size from 1 to 100 kDa, and are ribosomally synthesized. They can alter cell membrane integrity, can interfere with transcription, translation, or DNA replication, and frequently are produced during stationary phase. Some archaea do produce proteinaceous antibiotics. For example, Sulfolobus islandicus produces an...

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Halocins: salt-dependent bacteriocins produced by extremely halophilic rods

Cited by 94 publications

References 7 publications

Halocin S8: a 36-Amino-Acid Microhalocin from the Haloarchaeal Strain S8a

Halocin S8: a 36-Amino-Acid Microhalocin from the Haloarchaeal Strain S8a

Isolation, sequence, and expression of the gene encoding halocin H4, a bacteriocin from the halophilic archaeon Haloferax mediterranei R4

Secreted Euryarchaeal Microhalocins Kill Hyperthermophilic Crenarchaea

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