Thermal stability of a mercuric reductase from the Red Sea Atlantis II hot brine environment as analyzed by site-directed mutagenesis

Simple item page
dc.AffiliationOctober University for modern sciences and Arts (MSA)
dc.contributor.authorMaged M.
dc.contributor.authorHosseiny A.E.
dc.contributor.authorSaadeldin M.K.
dc.contributor.authorAziz R.K.
dc.contributor.authorRamadan E.
dc.contributor.otherDepartment of Biology
dc.contributor.otherSchool of Sciences and Engineering
dc.contributor.otherThe American University in Cairo
dc.contributor.otherNew Cairo
dc.contributor.otherEgypt; Department of Microbiology and Immunology
dc.contributor.otherFaculty of Pharmacy
dc.contributor.otherCairo University
dc.contributor.otherCairo
dc.contributor.otherEgypt; Faculty of Pharmacy
dc.contributor.otherThe British University in Egypt (BUE)
dc.contributor.otherEl Shorouk
dc.contributor.otherEgypt; Science and Technology Research Center
dc.contributor.otherSchool of Sciences and Engineering
dc.contributor.otherThe American University in Cairo
dc.contributor.otherNew Cairo
dc.contributor.otherEgypt; Faculty of Biotechnology
dc.contributor.otherOctober University for Modern Sciences and Arts
dc.contributor.other6th October City
dc.contributor.otherCairo
dc.contributor.otherEgypt
dc.date.accessioned2020-01-09T20:40:41Z
dc.date.available2020-01-09T20:40:41Z
dc.date.issued2019
dc.descriptionScopus
dc.description.abstractThe lower convective layer (LCL) of the Atlantis II brine pool of the Red Sea is a unique environment in terms of high salinity, temperature, and high concentrations of heavy metals. Mercuric reductase enzymes functional in such extreme conditions could be considered a potential tool in the environmental detoxification of mercurial poisoning and might alleviate ecological hazards in the mining industry. Here, we constructed a mercuric reductase library from Atlantis II, from which we identified genes encoding two thermostable mercuric reductase (MerA) isoforms: one is halophilic (designated ATII-LCL) while the other is not (designated ATII-LCLNH). The ATII-LCL MerA has a short motif composed of four aspartic acids (4D414- 417) and two characteristic signature boxes that played a crucial role in its thermal stability. To further understand the mechanism behind the thermostability of the two studied enzymes, we mutated the isoform ATII-LCL-NH and found that the substitution of 2 aspartic acids (2D) at positions 415 and 416 enhanced the thermal stability, while other mutations had the opposite effect. The 2D mutant showed superior thermal tolerance, as it retained 81% of its activity after 10 min of incubation at 70�C. A three-dimensional structure prediction revealed newly formed salt bridges and H bonds in the 2D mutant compared to the parent molecule. To the best of our knowledge, this study is the first to rationally design a mercuric reductase with enhanced thermal stability, which we propose to have a strong potential in the bioremediation of mercurial poisoning. � 2019 American Society for Microbiology.en_US
dc.identifier.doihttps://doi.org/10.1128/AEM.02387-18
dc.identifier.doiPubMedID30446558
dc.identifier.issn992240
dc.identifier.otherhttps://doi.org/10.1128/AEM.02387-18
dc.identifier.otherPubMedID30446558
dc.identifier.urihttps://t.ly/52yxR
dc.language.isoEnglishen_US
dc.publisherAmerican Society for Microbiologyen_US
dc.relation.ispartofseriesApplied and Environmental Microbiology
dc.relation.ispartofseries85
dc.subjectAtlantis IIen_US
dc.subjectBioprospectingen_US
dc.subjectBrine poolsen_US
dc.subjectExtreme environmentsen_US
dc.subjectMerAen_US
dc.subjectMercuric reductaseen_US
dc.subjectProtein engineeringen_US
dc.subjectRed Seaen_US
dc.subjectSite-directed mutagenesisen_US
dc.subjectThermostableen_US
dc.subjectAmino acidsen_US
dc.subjectBioremediationen_US
dc.subjectChemical bondsen_US
dc.subjectDetoxificationen_US
dc.subjectEnzymesen_US
dc.subjectHeavy metalsen_US
dc.subjectMutagenesisen_US
dc.subjectStabilityen_US
dc.subjectAtlantisen_US
dc.subjectBioprospectingen_US
dc.subjectBrine poolsen_US
dc.subjectExtreme environmenten_US
dc.subjectMerAen_US
dc.subjectMercuric reductaseen_US
dc.subjectProtein engineeringen_US
dc.subjectRed seaen_US
dc.subjectSite directed mutagenesisen_US
dc.subjectThermostableen_US
dc.subjectThermodynamic stabilityen_US
dc.subjectbioremediationen_US
dc.subjectbrineen_US
dc.subjectdetoxificationen_US
dc.subjectenzymeen_US
dc.subjectenzyme activityen_US
dc.subjectgenetic analysisen_US
dc.subjectheavy metalen_US
dc.subjecttemperature toleranceen_US
dc.subjectIndian Oceanen_US
dc.subjectRed Sea [Indian Ocean]en_US
dc.subjectbacterial proteinen_US
dc.subjectmercuric reductaseen_US
dc.subjectmercuryen_US
dc.subjectoxidoreductaseen_US
dc.subjectsea wateren_US
dc.subjectamino acid sequenceen_US
dc.subjectbacteriumen_US
dc.subjectchemistryen_US
dc.subjectecosystemen_US
dc.subjectenzyme stabilityen_US
dc.subjectenzymologyen_US
dc.subjectgeneticsen_US
dc.subjectheaten_US
dc.subjectIndian Oceanen_US
dc.subjectisolation and purificationen_US
dc.subjectkineticsen_US
dc.subjectmetabolismen_US
dc.subjectmicrobiologyen_US
dc.subjectprotein motifen_US
dc.subjectsequence alignmenten_US
dc.subjectsite directed mutagenesisen_US
dc.subjectAmino Acid Motifsen_US
dc.subjectAmino Acid Sequenceen_US
dc.subjectBacteriaen_US
dc.subjectBacterial Proteinsen_US
dc.subjectEcosystemen_US
dc.subjectEnzyme Stabilityen_US
dc.subjectHot Temperatureen_US
dc.subjectIndian Oceanen_US
dc.subjectKineticsen_US
dc.subjectMercuryen_US
dc.subjectMutagenesis, Site-Directeden_US
dc.subjectOxidoreductasesen_US
dc.subjectSeawateren_US
dc.subjectSequence Alignmenten_US
dc.titleThermal stability of a mercuric reductase from the Red Sea Atlantis II hot brine environment as analyzed by site-directed mutagenesisen_US
dc.typeArticleen_US
dcterms.isReferencedBy(2015) Red Sea. New World Encyclopedia, , http://www.newworldencyclopedia.org/p/index.php?title=Red_Sea&oldid=989212; Bower, A., (2009) R/V Oceanus Voyage 449-6 Red Sea Atlantis II Deep Complex Area, , http://www.whoi.edu/science/PO/people/www-abower/abower/techreps/KAUST_CTR0901_press.pdf, 19 October-1 November 2008. Woods Hole Oceanographic Institute Technical Report. Woods Hole Oceanographic Institute (WHOI) and King Abdullah University of Science and Technology, Woods Hole, MA; Botz, R., Schmidt, M., Wehner, H., Hufnagel, H., Stoffers, P., Organic-rich sediments in brine-filled Shaban-and Kebrit deeps, northern Red Sea (2007) Chem Geol, 244, pp. 520-553. , https://doi.org/10.1016/j.chemgeo.2007.07.004; Speth, D.R., Lagkouvardos, I., Wang, Y., Qian, P.-Y., Dutilh, B.E., Jetten, M.S.M., Draft genome of Scalindua rubra, obtained from the interface above the Discovery Deep brine in the Red Sea, sheds light on potential salt adaptation strategies in Anammox bacteria (2017) Microb Ecol, 74, pp. 1-5. , https://doi.org/10.1007/s00248-017-0929-7; Wang, Y., Cao, H., Zhang, G., Bougouffa, S., Lee, O.O., Al-Suwailem, A., Qian, P.Y., Autotrophic microbe metagenomes and metabolic pathways differentiate adjacent Red Sea brine pools (2013) Sci Rep, 3, p. 1748. , https://doi.org/10.1038/srep01748; Hartmann, M., Scholten, J.C., Stoffers, P., Wehner, F., Hydrographic structure of brine-filled deeps in the Red Sea-new results from the Shaban, Kebrit, Atlantis II, and Discovery Deep (1998) Mar Geol, 144, pp. 311-330. , https://doi.org/10.1016/S0025-3227(97)00055-8; Wang, Y., Yang, J., Lee, O.O., Dash, S., Lau, S.C., Al-Suwailem, A., Wong, T.Y., Qian, P.Y., Hydrothermally generated aromatic compounds are consumed by bacteria colonizing in Atlantis II Deep of the Red Sea (2011) ISME J, 5, pp. 1652-1659. , https://doi.org/10.1038/ismej.2011.42; Wang, Y., Li, J.T., He, L.S., Yang, B., Gao, Z.M., Cao, H.L., Batang, Z., Qian, P.Y., Zonation of microbial communities by a hydrothermal mound in the Atlantis II Deep (the Red Sea) (2015) PLoS One, 10. , https://doi.org/10.1371/journal.pone.0140766; Laurila, T.E., Hannington, M.D., Leybourne, M., Petersen, S., Devey, C.W., Garbe-Sch�nberg, D., New insights into the mineralogy of the Atlantis II Deep metalliferous sediments, Red Sea (2015) Geochem Geophys Geosyst, 16, pp. 4449-4478. , https://doi.org/10.1002/2015GC006010; Antunes, A., Ngugi, D.K., Stingl, U., Microbiology of the Red Sea (and other) deep-sea anoxic brine lakes (2011) Environ Microbiol Rep, 3, pp. 416-433. , https://doi.org/10.1111/j.1758-2229.2011.00264.x; Siam, R., Mustafa, G.A., Sharaf, H., Moustafa, A., Ramadan, A.R., Antunes, A., Bajic, V.B., Dorry, H.E., Unique prokaryotic consortia in geochemically distinct sediments from Red Sea Atlantis II and discovery deep brine pools (2012) PLoS One, 7. , https://doi.org/10.1371/journal.pone.0042872; Nascimento, A.M., Chartone-Souza, E., Operon mer: bacterial resistance to mercury and potential for bioremediation of contaminated environments (2003) Genet Mol Res, 2, pp. 92-101; Mathema, V.B., Thakuri, B.C., Sillanpaa, M., Bacterial mer operonmediated detoxification of mercurial compounds: a short review (2011) Arch Microbiol, 193, pp. 837-844. , https://doi.org/10.1007/s00203-011-0751-4; Moller, A.K., Barkay, T., Hansen, M.A., Norman, A., Hansen, L.H., Sorensen, S.J., Boyd, E.S., Kroer, N., Mercuric reductase genes (merA) and mercury resistance plasmids in High Arctic snow, freshwater and sea-ice brine (2014) FEMS Microbiol Ecol, 87, pp. 52-63. , https://doi.org/10.1111/1574-6941.12189; Zheng, R., Wu, S., Ma, N., Sun, C., Genetic and physiological adaptations of marine bacterium Pseudomonas stutzeri 273 to mercury stress (2018) Front Microbiol, 9, p. 682. , https://doi.org/10.3389/fmicb.2018.00682; Varekamp, J.C., Buseck, P.R., The speciation of mercury in hydrothermal systems, with applications to ore deposition (1984) Geochim Cosmochim Acta, 48, pp. 177-185. , https://doi.org/10.1016/0016-7037(84)90359-4; Gworek, B., Bemowska-Kalabun, O., Kijen?ka, M., Wrzosek-Jakubowska, J., Mercury in marine and oceanic waters-a review (2016) Water Air Soil Pollut, 227, p. 371. , https://doi.org/10.1007/s11270-016-3060-3; Ay, N., Content of mercury in some natural waters (1962) Chem Abstr, 57, p. 16336; Pempkowiak, J., Cossa, D., Sikora, A., Sanjuan, J., Mercury in water and sediments of the southern Baltic sea (1998) Sci Total Environ, 213, pp. 185-192; Ci, Z., Zhang, X., Wang, Z., Niu, Z., Phase speciation of mercury (Hg) in coastal water of the Yellow Sea, China (2011) Mar Chem, 126, pp. 250-255. , https://doi.org/10.1016/j.marchem.2011.06.004; Brown, N.L., Ford, S.J., Pridmore, R.D., Fritzinger, D.C., Nucleotide se-quence of a gene from the Pseudomonas transposon Tn501 encoding mercuric reductase (1983) Biochemistry, 22, pp. 4089-4095; Schiering, N., Kabsch, W., Moore, M.J., Distefano, M.D., Walsh, C.T., Pai, E.F., Structure of the detoxification catalyst mercuric ion reductase from Bacillus sp. strain RC607 (1991) Nature, 352, pp. 168-172. , https://doi.org/10.1038/352168a0; Miller, S.M., Moore, M.J., Massey, V., Williams, C.H., Jr., Distefano, M.D., Ballou, D.P., Walsh, C.T., Evidence for the participation of Cys558 and Cys559 at the active site of mercuric reductase (1989) Biochemistry, 28, pp. 1194-1205; Wang, Y., Freedman, Z., Lu-Irving, P., Kaletsky, R., Barkay, T., An initial characterization of the mercury resistance (mer) system of the thermophilic bacterium Thermus thermophilus HB27 (2009) FEMS Microbiol Ecol, 67, pp. 118-129. , https://doi.org/10.1111/j.1574-6941.2008.00603.x; Li, W.F., Zhou, X.X., Lu, P., Structural features of thermozymes (2005) Biotechnol Adv, 23, pp. 271-281. , https://doi.org/10.1016/j.biotechadv.2005.01.002; Vieille, C., Burdette, D.S., Zeikus, J.G., Thermozymes (1996) Biotechnol Annu Rev, 2, pp. 1-83; Huang, R., Yang, Q., Feng, H., Single amino acid mutation alters thermostability of the alkaline protease from Bacillus pumilus: thermodynamics and temperature dependence (2015) Acta Biochim Biophys Sin (Shanghai), 47, pp. 98-105. , https://doi.org/10.1093/abbs/gmu120; Trivedi, S., Gehlot, H.S., Rao, S.R., Protein thermostability in Archaea and Eubacteria (2006) Genet Mol Res, 5, pp. 816-827; Suzuki, Y., A general principle of increasing protein thermostability (1989) Proc Jpn Acad Ser B Phys Biol Sci, 65, pp. 146-148; Sayed, A., Ghazy, M.A., Ferreira, A.J., Setubal, J.C., Chambergo, F.S., Ouf, A., Adel, M., El-Dorry, H., A novel mercuric reductase from the unique deep brine environment of Atlantis II in the Red Sea (2014) J Biol Chem, 289, pp. 1675-1687. , https://doi.org/10.1074/jbc.M113.493429; Artz, J.H., White, S.N., Zadvornyy, O.A., Fugate, C.J., Hicks, D., Gauss, G.H., Posewitz, M.C., Peters, J.W., Biochemical and structural properties of a thermostable mercuric ion reductase from Metallosphaera sedula (2015) Front Bioeng Biotechnol, 3, p. 97. , https://doi.org/10.3389/fbioe.2015.00097; Bafana, A., Khan, F., Suguna, K., Structural and functional characterization of mercuric reductase from Lysinibacillus sphaericus strain G1 (2017) Biometals, 30, pp. 809-819. , https://doi.org/10.1007/s10534-017-0050-x; Baker, E.N., Hubbard, R.E., Hydrogen bonding in globular proteins (1984) Prog Biophys Mol Biol, 44, pp. 97-179; Petsko, G.A., Ringe, D., (2004) Bonds that stabilize folded proteins, p 10-11, , Protein structure and function. New Science Press, London, United Kingdom; Costantini, S., Colonna, G., Facchiano, A.M., ESBRI: a web server for evaluating salt bridges in proteins (2008) Bioinformation, 3, pp. 137-138; Celniker, G., Nimrod, G., Ashkenazy, H., Glaser, F., Martz, E., Mayrose, I., Pupko, T., Ben-Tal, N., ConSurf: using evolutionary data to raise testable hypotheses about protein function (2013) Isr J Chem, 53, pp. 199-206. , https://doi.org/10.1002/ijch.201200096; Madern, D., Ebel, C., Zaccai, G., Halophilic adaptation of enzymes (2000) Extremophiles, 4, pp. 91-98; Mevarech, M., Frolow, F., Gloss, L.M., Halophilic enzymes: proteins with a grain of salt (2000) Biophys Chem, 86, pp. 155-164. , https://doi.org/10.1016/S0301-4622(00)00126-5; Fukuchi, S., Yoshimune, K., Wakayama, M., Moriguchi, M., Nishikawa, K., Unique amino acid composition of proteins in halophilic bacteria (2003) J Mol Biol, 327, pp. 347-357. , https://doi.org/10.1016/S0022-2836(03)00150-5; Siglioccolo, A., Paiardini, A., Piscitelli, M., Pascarella, S., Structural adaptation of extreme halophilic proteins through decrease of conserved hydrophobic contact surface (2011) BMC Struct Biol, 11, p. 50. , https://doi.org/10.1186/1472-6807-11-50; Zeroual, Y.M.A., Dzairi, F.Z., Talbi, M., Chung, P.U., Lee, K., Purification and characterization of cytosolic mercuric reductase from Klebsiella pneumoniae (2003) Ann Microbiol, 53, pp. 149-160; Takano, K., Higashi, R., Okada, J., Mukaiyama, A., Tadokoro, T., Koga, Y., Kanaya, S., Proline effect on the thermostability and slow unfolding of a hyperthermophilic protein (2009) J Biochem, 145, pp. 79-85. , https://doi.org/10.1093/jb/mvn144; Watanabe, K., Masuda, T., Ohashi, H., Mihara, H., Suzuki, Y., Multiple proline substitutions cumulatively thermostabilize Bacillus cereus ATCC7064 oligo-1,6-glucosidase Irrefragable proof supporting the proline rule (1994) Eur J Biochem, 226, pp. 277-283; Watanabe, K., Kitamura, K., Suzuki, Y., Analysis of the critical sites for protein thermostabilization by proline substitution in oligo-1,6-glucosidase from Bacillus coagulans ATCC 7050 and the evolutionary consideration of proline residues (1996) Appl Environ Microbiol, 62, pp. 2066-2073; Hardy, F., Vriend, G., Veltman, O.R., van der Vinne, B., Venema, G., Eijsink, V.G., Stabilization of Bacillus stearothermophilus neutral protease by introduction of prolines (1993) FEBS Lett, 317, pp. 89-92. , https://doi.org/10.1016/0014-5793(93)81497-N; Imanaka, T., Molecular bases of thermophily in hyperthermophiles (2011) Proc Jpn Acad Ser B Phys Biol Sci, 87, pp. 587-602. , https://doi.org/10.2183/pjab.87.587; Kumar, S., Ma, B., Tsai, C.J., Nussinov, R., Electrostatic strengths of salt bridges in thermophilic and mesophilic glutamate dehydrogenase monomers (2000) Proteins, 38, pp. 368-383; Xu, D., Tsai, C.J., Nussinov, R., Hydrogen bonds and salt bridges across protein-protein interfaces (1997) Protein Eng, 10, pp. 999-1012; Kamanda Ngugi, D., Blom, J., Alam, I., Rashid, M., Ba-Alawi, W., Zhang, G., Hikmawan, T., Stingl, U., Comparative genomics reveals adaptations of a halotolerant thaumarchaeon in the interfaces of brine pools in the Red Sea (2015) ISME J, 9, pp. 396-411. , https://doi.org/10.1038/ismej.2014.137; Sonbol, S.A., Ferreira, A.J., Siam, R., Red Sea Atlantis II brine pool nitrilase with unique thermostability profile and heavy metal tolerance (2016) BMC Biotechnol, 16, p. 14. , https://doi.org/10.1186/s12896-016-0244-2; Corpet, F., Multiple sequence alignment with hierarchical clustering (1988) Nucleic Acids Res, 16, pp. 10881-10890; Mount, D.W., Using the Basic Local Alignment Search Tool (BLAST) (2007) CSH Protoc, 2007. , https://doi.org/10.1101/pdb.top17; Kalmbach, S., Manz, W., Wecke, J., Szewzyk, U., Aquabacterium gen. nov., with description of Aquabacterium citratiphilum sp. nov., Aquabacterium parvum sp. nov. and Aquabacterium commune sp. nov., three in situ dominant bacterial species from the Berlin drinking water system (1999) Int J Syst Bacteriol, 49, pp. 769-777. , https://doi.org/10.1099/00207713-49-2-769; Fox, B., Walsh, C.T., Mercuric reductase Purification and characterization of a transposon-encoded flavoprotein containing an oxidationreduction-active disulfide (1982) J Biol Chem, 257, pp. 2498-2503; Ledwidge, R., Hong, B., Dotsch, V., Miller, S.M., NmerA of Tn501 mercuric ion reductase: structural modulation of the pKa values of the metal binding cysteine thiols (2010) Biochemistry, 49, pp. 8988-8998. , https://doi.org/10.1021/bi100537f; Ledwidge, R., Patel, B., Dong, A., Fiedler, D., Falkowski, M., Zelikova, J., Summers, A.O., Miller, S.M., NmerA, the metal binding domain of mercuric ion reductase, removes Hg2 + from proteins, delivers it to the catalytic core, and protects cells under glutathione-depleted conditions (2005) Biochemistry, 44, pp. 11402-11416. , https://doi.org/10.1021/bi050519d; Biasini, M., Bienert, S., Waterhouse, A., Arnold, K., Studer, G., Schmidt, T., Kiefer, F., Schwede, T., SWISSMODEL: modelling protein tertiary and quaternary structure using evolutionary information (2014) Nucleic Acids Res, 42, pp. W252-W258. , https://doi.org/10.1093/nar/gku340; Holec, P.V., Hackel, B.J., PyMOL360: multi-user gamepad control of molecular visualization software (2016) J Comput Chem, 37, pp. 2667-2669. , https://doi.org/10.1002/jcc.24489; Sarakatsannis, J.N., Duan, Y., Statistical characterization of salt bridges in proteins (2005) Proteins, 60, pp. 732-739. , https://doi.org/10.1002/prot.20549; Kumar, S., Nussinov, R., Relationship between ion pair geometries and electrostatic strengths in proteins (2002) Biophys J, 83, pp. 1595-1612. , https://doi.org/10.1016/S0006-3495(02)73929-5; Zhang, Y., Bond, C.S., Bailey, S., Cunningham, M.L., Fairlamb, A.H., Hunter, W.N., The crystal structure of trypanothione reductase from the human pathogen Trypanosoma cruzi at 2.3 � resolution (1996) Protein Sci, 5, pp. 52-61. , https://doi.org/10.1002/pro.5560050107
dcterms.sourceScopus

Files

Original bundle

Now showing 1 - 2 of 2
Thumbnail Image
Name:
avatar_scholar_256.png
Size:
6.31 KB
Format:
Portable Network Graphics
Description:
Download
Thumbnail Image
Name:
AEM.02387-18.pdf
Size:
1.07 MB
Format:
Adobe Portable Document Format
Description:
Download

Collections

Faculty of Biotechnology Research Paper