Characterization of the PH1704 Protease from Pyrococcus horikoshii OT3 and the Critical Functions of Tyr120

Zhan, Daqian; Bai, Aixi; Yu, Lei; Han, Wei; Feng, Yan

doi:10.1371/journal.pone.0103902

Cited by 10 publications

(9 citation statements)

References 36 publications

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“…These enzymes represent a large percentage of the global enzyme market [ 12 , 13 ]. Most proteases from extremophiles belong to the serine type and many of them come from hyperthermophilic archaea belonging to the genera Pyrococcus [ 14 ], Thermococcus [ 15 ], Desulfurococcus [ 16 ], Pyrobaculum [ 17 ], Staphylothermus [ 18 ], and from the thermoacidophilic archaeon Sulfolobus [ 19 ] (see Table 3 ).…”

Section: Proteolytic Enzymes (Ec 34xx)mentioning

confidence: 99%

Biotechnological applications of archaeal enzymes from extreme environments

Cabrera

Blamey

2018

Biol Res

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To date, many industrial processes are performed using chemical compounds, which are harmful to nature. An alternative to overcome this problem is biocatalysis, which uses whole cells or enzymes to carry out chemical reactions in an environmentally friendly manner. Enzymes can be used as biocatalyst in food and feed, pharmaceutical, textile, detergent and beverage industries, among others. Since industrial processes require harsh reaction conditions to be performed, these enzymes must possess several characteristics that make them suitable for this purpose. Currently the best option is to use enzymes from extremophilic microorganisms, particularly archaea because of their special characteristics, such as stability to elevated temperatures, extremes of pH, organic solvents, and high ionic strength. Extremozymes, are being used in biotechnological industry and improved through modern technologies, such as protein engineering for best performance. Despite the wide distribution of archaea, exist only few reports about these microorganisms isolated from Antarctica and very little is known about thermophilic or hyperthermophilic archaeal enzymes particularly from Antarctica. This review summarizes current knowledge of archaeal enzymes with biotechnological applications, including two extremozymes from Antarctic archaea with potential industrial use, which are being studied in our laboratory. Both enzymes have been discovered through conventional screening and genome sequencing, respectively.

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Section: Proteolytic Enzymes (Ec 34xx)mentioning

confidence: 99%

Biotechnological applications of archaeal enzymes from extreme environments

Cabrera

Blamey

2018

Biol Res

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“…Lys43 is also in the immediate neighborhood of the active site, the involvement of which has not been determined; it has been implicated in substrate selection. Tyr120, which was deemed to be important in accessibility to the active site in the P. horikoshii protease (Zhan et al, 2014), is however not present in the protease from T. thioreducens. At position 120 there is a threonine.…”

Section: Resultsmentioning

confidence: 99%

“…Subsequent biochemical investigations have indicated that in addition to the catalytic triad, Lys48 and especially Tyr120 are important residues in substrate selection and catalysis. The latter residue has been described as a 'gateway' to the active site (Zhan et al, 2014).…”

Section: Introductionmentioning

confidence: 99%

The structure of the Pfp1 protease from the hyperthermophilic archaeonThermococcus thioreducensin two crystal forms

Larson

McPherson

2017

Acta Cryst Sect D Struct Biol

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The Pfp1 protease, a cysteine protease of unknown specificity from the hyperthermophilic archaeon Thermococcus thioreducens, was crystallized in two distinctive crystal forms: from concentrated citrate in one case and PEG in the other. X-ray data were collected from both crystal forms at room temperature to about 1.9 Å resolution using a laboratory source and detector, and the structures were solved by molecular replacement using the Pfp1 protease from Pyrococcus horikoshii as the search model. In the T. thioreducens protease structures, Cys18 residues on adjacent molecules in the asymmetric units form intermolecular disulfide bonds, thereby yielding hexamers composed of three cross-linked, quasi-dyad-related dimers with crystallographically exact threefold axes and exhibiting almost exact 32 symmetry. The corresponding residue in P. horikoshii Pfp1 is Tyr18. An individual active site containing Cys100 and His101 also includes a Glu74 residue contributed by a quasi-twofold-related, non-cross-linked subunit. Two catalytic triads are therefore closely juxtaposed about the quasi-twofold axis at the interface of these subunits, and are relatively sequestered within the hexamer cavity. The cysteine in the active site is observed to be oxidized in both of the crystal forms that were studied.

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“…A Glu residue, instead of an Asp, completes the catalytic triad (Figs 1b and 2b ). Class-III orthologs PH1704, a protease from Pyrococcus horikoshii [ 39 ], I3RG07, an intracellular protease from Pyrococcus sp . ST04 and and PfpI [ 40 ], an intracellular protease from the hyperthermophilic Sulfolobus solfataricus are devoid of P-domain and the ‘linker region’ ( Fig 2c ).…”

Section: Resultsmentioning

confidence: 99%

Structural and biochemical studies on Vibrio cholerae Hsp31 reveals a novel dimeric form and Glutathione-independent Glyoxalase activity

et al. 2017

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Vibrio cholerae experiences a highly hostile environment at human intestine which triggers the induction of various heat shock genes. The hchA gene product of V. cholerae O395, referred to a hypothetical intracellular protease/amidase VcHsp31, is one such stress-inducible homodimeric protein. Our current study demonstrates that VcHsp31 is endowed with molecular chaperone, amidopeptidase and robust methylglyoxalase activities. Through site directed mutagenesis coupled with biochemical assays on VcHsp31, we have confirmed the role of residues in the vicinity of the active site towards amidopeptidase and methylglyoxalase activities. VcHsp31 suppresses the aggregation of insulin in vitro in a dose dependent manner. Through crystal structures of VcHsp31 and its mutants, grown at various temperatures, we demonstrate that VcHsp31 acquires two (Type-I and Type-II) dimeric forms. Type-I dimer is similar to EcHsp31 where two VcHsp31 monomers associate in eclipsed manner through several intersubunit hydrogen bonds involving their P-domains. Type-II dimer is a novel dimeric organization, where some of the intersubunit hydrogen bonds are abrogated and each monomer swings out in the opposite directions centering at their P-domains, like twisting of wet cloth. Normal mode analysis (NMA) of Type-I dimer shows similar movement of the individual monomers. Upon swinging, a dimeric surface of ~400Å2, mostly hydrophobic in nature, is uncovered which might bind partially unfolded protein substrates. We propose that, in solution, VcHsp31 remains as an equilibrium mixture of both the dimers. With increase in temperature, transformation to Type-II form having more exposed hydrophobic surface, occurs progressively accounting for the temperature dependent increase of chaperone activity of VcHsp31.

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Characterization of the PH1704 Protease from Pyrococcus horikoshii OT3 and the Critical Functions of Tyr120

Cited by 10 publications

References 36 publications

Biotechnological applications of archaeal enzymes from extreme environments

Biotechnological applications of archaeal enzymes from extreme environments

The structure of the Pfp1 protease from the hyperthermophilic archaeonThermococcus thioreducensin two crystal forms

Structural and biochemical studies on Vibrio cholerae Hsp31 reveals a novel dimeric form and Glutathione-independent Glyoxalase activity

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