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The effects of an engineered cation site on the structure, activity, and EPR properties of cytochrome c peroxidase.

Bonagura CA, Sundaramoorthy M, Bhaskar B, Poulos TL
Biochemistry 38 5538-45 (1999)
Cited: 19 times
EuropePMC logo PMID: 10220341

Abstract

Earlier work [Bonagura et al. (1996) Biochemistry 35, 6107] showed that the K+ site found in the proximal pocket of ascorbate peroxidase (APX) could be engineered into cytochrome c peroxidase (CCP). Binding of K+ at the engineered site results in a loss in activity and destabilization of the CCP compound I Trp191 cationic radical owing to long-range electrostatic effects. The engineered CCP mutant crystal structure has been refined to 1.5 A using data obtained at cryogenic temperatures which provides a more detailed basis for comparison with the naturally occurring K+ site in APX. The characteristic EPR signal associated with the Trp191 radical becomes progressively weaker as K+ is added, which correlates well with the loss in enzyme activity as [K+] is increased. These results coupled with stopped-flow studies support our earlier conclusions that the loss in activity and EPR signal is due to destabilization of the Trp191 cationic radical.

Reviews citing this publication (4)

  1. Heme enzyme structure and function. Poulos TL. Chem Rev 114 3919-3962 (2014)
  2. Mechanisms of compound I formation in heme peroxidases. Hiner AN, Raven EL, Thorneley RN, García-Cánovas F, Rodríguez-López JN. J Inorg Biochem 91 27-34 (2002)
  3. Tailoring new enzyme functions by rational redesign. Cedrone F, Ménez A, Quéméneur E. Curr Opin Struct Biol 10 405-410 (2000)
  4. The role of the heme propionates in heme biochemistry. Guallar V, Olsen B. J Inorg Biochem 100 755-760 (2006)

Articles citing this publication (15)

  1. Crystal structure of maltose phosphorylase from Lactobacillus brevis: unexpected evolutionary relationship with glucoamylases. Egloff MP, Uppenberg J, Haalck L, van Tilbeurgh H. Structure 9 689-697 (2001)
  2. Artificial protein cavities as specific ligand-binding templates: characterization of an engineered heterocyclic cation-binding site that preserves the evolved specificity of the parent protein. Musah RA, Jensen GM, Bunte SW, Rosenfeld RJ, Goodin DB. J Mol Biol 315 845-857 (2002)
  3. Detection of a tryptophan radical in the reaction of ascorbate peroxidase with hydrogen peroxide. Hiner AN, Martínez JI, Arnao MB, Acosta M, Turner DD, Lloyd Raven E, Rodríguez-López JN. Eur J Biochem 268 3091-3098 (2001)
  4. Crystal structure of Leishmania major peroxidase and characterization of the compound i tryptophan radical. Jasion VS, Polanco JA, Meharenna YT, Li H, Poulos TL. J Biol Chem 286 24608-24615 (2011)
  5. Substrate specificity of lignin peroxidase and a S168W variant of manganese peroxidase. Timofeevski SL, Nie G, Reading NS, Aust SD. Arch Biochem Biophys 373 147-153 (2000)
  6. Crystal structure of the Leishmania major peroxidase-cytochrome c complex. Jasion VS, Doukov T, Pineda SH, Li H, Poulos TL. Proc Natl Acad Sci U S A 109 18390-18394 (2012)
  7. Introduction and characterization of a functionally linked metal ion binding site at the exposed heme edge of myoglobin. Hunter CL, Maurus R, Mauk MR, Lee H, Raven EL, Tong H, Nguyen N, Smith M, Brayer GD, Mauk AG. Proc Natl Acad Sci U S A 100 3647-3652 (2003)
  8. Phylogenetic analysis, molecular modeling, substrate-inhibitor specificity, and active site comparison of bacterial, fungal, and plant heme peroxidases. Singh S, Pandey VP, Naaz H, Dwivedi UN. Biotechnol Appl Biochem 59 283-294 (2012)
  9. Using an artificial tryptophan "wire" in cytochrome c peroxidase for oxidation of organic substrates. Field MJ, Bains RK, Warren JJ. Dalton Trans 46 11078-11083 (2017)
  10. Binding of imidazole, 1-methylimidazole and 4-nitroimidazole to yeast cytochrome c peroxidase (CcP) and the distal histidine mutant, CcP(H52L). Erman JE, Chinchilla D, Studer J, Vitello LB. Biochim Biophys Acta 1854 869-881 (2015)
  11. Role of proximal methionine residues in Leishmania major peroxidase. Yadav RK, Pal S, Dolai S, Adak S. Arch Biochem Biophys 515 21-27 (2011)
  12. Trapping of peptide-based surrogates in an artificially created channel of cytochrome c peroxidase. Hays AM, Gray HB, Goodin DB. Protein Sci 12 278-287 (2003)
  13. Why Do Most Aromatics Fail to Support Hole Hopping in the Cytochrome c Peroxidase-Cytochrome c Complex? Ru X, Crane BR, Zhang P, Beratan DN. J Phys Chem B 125 7763-7773 (2021)
  14. Computational analysis of the tryptophan cation radical energetics in peroxidase Compound I. Poulos TL, Kim JS, Murarka VC. J Biol Inorg Chem 27 229-237 (2022)
  15. Crystal structure of Trypanosoma cruzi heme peroxidase and characterization of its substrate specificity and compound I intermediate. Freeman SL, Skafar V, Kwon H, Fielding AJ, Moody PCE, Martínez A, Issoglio FM, Inchausti L, Smircich P, Zeida A, Piacenza L, Radi R, Raven EL. J Biol Chem 298 102204 (2022)


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