Haloarchaeal proteases and proteolytic systems

Castro, Rosana Esther de; Maupin-Furlow, Julie A.; Giménez, María Inés; Seitz, María Karina Herrera; Sánchez, Jorge

doi:10.1111/j.1574-6976.2005.00003.x

Cited by 63 publications

(46 citation statements)

References 149 publications

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“…Therefore, due to their ability to tolerate conditions which inactivate their mesophilic counterparts, their study is of interest for both basic and applied research. Extracellular proteases have been puriWed and characterized at the biochemical and molecular levels from haloarchaea [1]. Scarce information is available on haloalkaliphilic archaea (optimum growth in 4-5 M NaCl, pH 8.5-12) and a few enzymes, including proteases, have been characterized from this group [1,2,6].…”

Section: Introductionmentioning

confidence: 99%

Effect of organic solvents on the activity and stability of an extracellular protease secreted by the haloalkaliphilic archaeon Natrialba magadii

Ruiz

Castro

2006

J Ind Microbiol Biotechnol

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The effect of various organic solvents on the activity and stability of an extracellular protease produced by the haloalkaliphilic archaeon Natrialba magadii was tested. This protease was active and stable in aqueous-organic solvent mixtures containing 1.5 M NaCl and glycerol, dimethylsulfoxide (DMSO), N,N-dimethyl formamide, propylenglycol, and dioxane. Among the solvents tested, DMSO, propylenglycol, and glycerol were effective in preserving enzyme stability in suboptimal NaCl concentrations. The stabilizing effect of DMSO on this haloalkaliphilic protease was more efficient at pH 8 than at pH 10, suggesting that DMSO may not substitute for salt to allow halophilic proteins to withstand the effect of high pH values. These results show that Nab. magadii extracellular protease is a solvent tolerant enzyme and suggest a potential application of this haloalkaliphilic protease in aqueous-organic solvent biocatalysis.

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Section: Introductionmentioning

confidence: 99%

Effect of organic solvents on the activity and stability of an extracellular protease secreted by the haloalkaliphilic archaeon Natrialba magadii

Ruiz

Castro

2006

J Ind Microbiol Biotechnol

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“…Almost all of the extracellular proteases isolated from haloarchaea are subtilisin-like serine proteases (subtilases) (2), known as halolysins (1,3). With whole-genome sequence data for many organisms quickly becoming available, an increasing number of halolysin genes have been identified, emphasizing the physiological significance of halolysins in the haloarchaea.…”

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“…With whole-genome sequence data for many organisms quickly becoming available, an increasing number of halolysin genes have been identified, emphasizing the physiological significance of halolysins in the haloarchaea. Because halolysins are active and stable under hypersaline conditions, they not only represent an attractive model for investigating the molecular basis of the halophilicity of enzymes but also show great potential as biocatalysts for the synthesis of oligopeptides due to the increase of their esterase/amidase activity ratios in organic mixtures of low water content (1,4,5). Additionally, the application of halophilic proteases at high salt concentrations can minimize the risk of microbial contamination (6).…”

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“…aloarchaea thrive in hypersaline environments such as solar salterns, salt lakes, and salt deposits, and many of them secrete proteases to degrade proteins into small peptides and amino acids which are then fed into central metabolism (1). Almost all of the extracellular proteases isolated from haloarchaea are subtilisin-like serine proteases (subtilases) (2), known as halolysins (1,3).…”

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Chitin Accelerates Activation of a Novel Haloarchaeal Serine Protease That Deproteinizes Chitin-Containing Biomass

Zhang

Wang

et al. 2014

Appl Environ Microbiol

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bThe haloarchaeon Natrinema sp. strain J7-2 has the ability to degrade chitin, and its genome harbors a chitin metabolism-related gene cluster that contains a halolysin gene, sptC. The sptC gene encodes a precursor composed of a signal peptide, an N-terminal propeptide consisting of a core domain (N*) and a linker peptide, a subtilisin-like catalytic domain, a polycystic kidney disease domain (PkdD), and a chitin-binding domain (ChBD). Here we report that the autocatalytic maturation of SptC is initiated by cis-processing of N* to yield an autoprocessed complex (N*-I WT ), followed by trans-processing/degradation of the linker peptide, the ChBD, and N*. The resulting mature form (M WT ) containing the catalytic domain and the PkdD showed optimum azocaseinolytic activity at 3 to 3.5 M NaCl, demonstrating salt-dependent stability. Deletion analysis revealed that the PkdD did not confer extra stability on the enzyme but did contribute to enzymatic activity. The ChBD exhibited salt-dependent chitinbinding capacity and mediated the binding of N*-I WT to chitin. ChBD-mediated chitin binding enhances SptC maturation by promoting activation of the autoprocessed complex. Our results also demonstrate that SptC is capable of removing proteins from shrimp shell powder (SSP) at high salt concentrations. Interestingly, N*-I WT released soluble peptides from SSP faster than did M WT . Most likely, ChBD-mediated binding of the autoprocessed complex to chitin in SSP not only accelerates enzyme activation but also facilitates the deproteinization process by increasing the local protease concentration around the substrate. By virtue of these properties, SptC is highly attractive for use in preparation of chitin from chitin-containing biomass.

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“…Most of these proteases belong to the subtilisin-like serine protease (subtilase) superfamily, known as halolysins (23,24). We previously cloned three halolysin-encoding genes from the genome of the haloarchaeon Natrinema sp.…”

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Alternative Translation Initiation of a Haloarchaeal Serine Protease Transcript Containing Two In-Frame Start Codons

Tang

et al. 2016

J Bacteriol

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Recent studies have shown that haloarchaea employ leaderless and Shine-Dalgarno (SD)-less mechanisms for translation initiation of leaderless transcripts with a 5= untranslated region (5= UTR) of <10 nucleotides (nt) and leadered transcripts with a 5= UTR of >10 nt, respectively. However, whether the two mechanisms can operate on the same naturally occurring haloarchaeal transcript carrying multiple potential start codons is unknown. In this study, the transcript of the sptA gene (encoding an extracellular serine protease of Natrinema sp. strain J7-2) was experimentally determined and found to contain two potential in-frame AUG codons (AUG 1 and AUG 2 ) located 5 and 29 nt, respectively, downstream of the transcription start site. Mutational analysis revealed that both AUGs can function as the translation start codon for production of active SptA, although AUG 1 is more efficient than AUG 2 for translation initiation. Insertion of a stable stem-loop structure between the two AUGs completely abolished initiation at AUG 1 but did not affect initiation at AUG 2 , indicating that AUG 2 -initiated translation does not involve ribosome scanning from the 5= end of the transcript. Furthermore, the efficiency of AUG 2 -initiated translation was not influenced by an upstream SD-like sequence. In addition, both AUG 1 and AUG 2 contribute to transcript stability, probably by recruiting ribosomes to protect the transcript against degradation. These data suggest that depending on which of two in-frame start codons is used, the sptA transcript can act as either a leaderless or a leadered transcript for SptA production in haloarchaea. IMPORTANCEIn eukaryotes and bacteria, alternative translation start sites contribute to proteome complexity and can be used as a functional mechanism to increase translation efficiency. However, little is known about alternative translation initiation in archaea. Our results demonstrate that leaderless and SD-less mechanisms can be used for translation initiation of the sptA transcript from two in-frame start codons, raising the possibility that in haloarchaea, alternative translation initiation on one transcript functions to increase translation efficiency and/or contribute to proteome complexity.

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Haloarchaeal proteases and proteolytic systems

Cited by 63 publications

References 149 publications

Effect of organic solvents on the activity and stability of an extracellular protease secreted by the haloalkaliphilic archaeon Natrialba magadii

Effect of organic solvents on the activity and stability of an extracellular protease secreted by the haloalkaliphilic archaeon Natrialba magadii

Chitin Accelerates Activation of a Novel Haloarchaeal Serine Protease That Deproteinizes Chitin-Containing Biomass

Alternative Translation Initiation of a Haloarchaeal Serine Protease Transcript Containing Two In-Frame Start Codons

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