Enhancement of hydrogen peroxide stability of a novel Anabaena sp. DyP-type peroxidase by site-directed mutagenesis of methionine residues

Ogola, Henry Joseph Oduor; Hashimoto, Naoya; Miyabe, Suguru; Ashida, Hiroyuki; Ishikawa, Takahiro; Shibata, Hitoshi; Sawa, Yousuke

doi:10.1007/s00253-010-2603-6

Cited by 40 publications

(28 citation statements)

References 33 publications

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“…On the contrary, only 18% dye degradation was observed. This effect is well reported and has been attributed to the fact that excessive H 2 O 2 can irreversibly oxidize/damage the enzyme and reduce its catalytic activity . In addition to emphasizing the importance of optimizing various parameters/reagent for efficient enzymatic reactions, this observation was used as an interesting discussion point with the students at the end of the experiment and to introduce the concepts of “oxyradicals” and “oxidants” and “antioxidants” for human health.…”

Section: Resultsmentioning

confidence: 99%

Raising environmental awareness through applied biochemistry laboratory experiments

Ashraf

2013

Biochem Molecular Bio Educ

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Our environment is under constant pressure and threat from various sources of pollution. Science students, in particular chemistry students, must not only be made aware of these issues, but also be taught that chemistry (and science) can provide solutions to such real-life issues. To this end, a newly developed biochemistry laboratory experiment is described that guides students to learn about the applicability of peroxidase enzymes to degrade organic dyes (as model pollutants) in simulated waste water. In addition to showing how enzymes can potentially be used for waste water remediation, various factors than can affect enzyme-based reactions such as pH, temperature, concentration of substrates/enzymes, and denaturants can also be tested. This "applied biotechnology" experiment was successfully implemented in an undergraduate biochemistry laboratory course to enhance students' learning of environmental issues as well important biochemistry concepts. Student survey confirmed that this laboratory experiment was successful in achieving the objectives of raising environmental awareness in students and illustrating the usefulness of chemistry in solving real-life problems. This experiment can be easily adopted in an introductory biochemistry laboratory course and taught as an inquiryguided exercise.

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Section: Resultsmentioning

confidence: 99%

Raising environmental awareness through applied biochemistry laboratory experiments

Ashraf

2013

Biochem Molecular Bio Educ

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“…That enhanced activity could be explained by the high turnover rate (k cat ϭ 419 s Ϫ1 ) ( (37) and Anabaena sp. DyP (24). On the other hand, an excess of the tested ions (2 mM) produced a slight decrease in enzyme activity, and only Fe 3ϩ , Hg 2ϩ , Pb 2ϩ , and NaN 3 severely affected enzyme performance.…”

Section: Discussionmentioning

confidence: 99%

Characterization of a Novel Dye-Decolorizing Peroxidase (DyP)-Type Enzyme from Irpex lacteus and Its Application in Enzymatic Hydrolysis of Wheat Straw

Salvachúa

Prieto

Martı́nez

et al. 2013

Appl Environ Microbiol

135

122

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Irpex lacteus is a white rot basidiomycete proposed for a wide spectrum of biotechnological applications which presents an interesting, but still scarcely known, enzymatic oxidative system. Among these enzymes, the production, purification, and identification of a new dye-decolorizing peroxidase (DyP)-type enzyme, as well as its physico-chemical, spectroscopic, and catalytic properties, are described in the current work. According to its N-terminal sequence and peptide mass fingerprinting analyses, I. lacteus DyP showed high homology (>95%) with the hypothetical (not isolated or characterized) protein cpop21 from an unidentified species of the family Polyporaceae. The enzyme had a low optimal pH, was very stable to acid pH and temperature, and showed improved activity and stability at high H 2 O 2 concentrations compared to other peroxidases. Other attractive features of I. lacteus DyP were its high catalytic efficiency oxidizing the recalcitrant anthraquinone and azo dyes assayed (k cat /K m of 1.6 ؋ 10 6 s ؊1 M ؊1 ) and its ability to oxidize nonphenolic aromatic compounds like veratryl alcohol. In addition, the effect of this DyP during the enzymatic hydrolysis of wheat straw was checked. The results suggest that I. lacteus DyP displayed a synergistic action with cellulases during the hydrolysis of wheat straw, increasing significantly the fermentable glucose recoveries from this substrate. These data show a promising biotechnological potential for this enzyme.

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“…Phe residues, in order to increase its resistance to hydrogen peroxide [221]. The heme cavity mutants M401L, M401I, M401F, and M451I had significantly increased hydrogen peroxide stabilities of 2.4-, 3.7-, 8.2-, and 5.2-fold, respectively.…”

Section: Site Directed Mutagenesismentioning

confidence: 99%

Hydrogen Peroxide in Biocatalysis. A Dangerous Liaison

Hernández

Berenguer-Murcia²,

Rodrigues³

et al. 2012

COC

140

107

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Hydrogen peroxide is a substrate or side-product in many enzyme-catalyzed reactions. For example, it is a side-product of oxidases, resulting from the re-oxidation of FAD with molecular oxygen, and it is a substrate for peroxidases and other enzymes. However, hydrogen peroxide is able to chemically modify the peptide core of the enzymes it interacts with, and also to produce the oxidation of some cofactors and prostetic groups (e.g., the hemo group). Thus, the development of strategies that may permit to increase the stability of enzymes in the presence of this deleterious reagent is an interesting target. This enhancement in enzyme stability has been attempted following almost all available strategies: site-directed mutagenesis (eliminating the most reactive moieties), medium engineering (using stabilizers), immobilization and chemical modification (trying to generate hydrophobic environments surrounding the enzyme, to confer higher rigidity to the protein or to generate oxidation-resistant groups), or the use of systems capable of decomposing hydrogen peroxide under very mild conditions. If hydrogen peroxide is just a side-product, its immediate removal has been reported to be the best solution. In some cases, when hydrogen peroxide is the substrate and its decomposition is not a sensible solution, researchers coupled one enzyme generating hydrogen peroxide "in situ" to the target enzyme resulting in a continuous supply of this reagent at low concentrations thus preventing enzyme inactivation.This review will focus on the general role of hydrogen peroxide in biocatalysis, the main mechanisms of enzyme inactivation produced by this reactive and the different strategies used to prevent enzyme inactivation caused by this "dangerous liaison". Outlook

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Enhancement of hydrogen peroxide stability of a novel Anabaena sp. DyP-type peroxidase by site-directed mutagenesis of methionine residues

Cited by 40 publications

References 33 publications

Raising environmental awareness through applied biochemistry laboratory experiments

Raising environmental awareness through applied biochemistry laboratory experiments

Characterization of a Novel Dye-Decolorizing Peroxidase (DyP)-Type Enzyme from Irpex lacteus and Its Application in Enzymatic Hydrolysis of Wheat Straw

Hydrogen Peroxide in Biocatalysis. A Dangerous Liaison

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