Structural Evidence of Substrate Specificity in Mammalian Peroxidases

Sheikh, Ishfaq Ahmed; Singh, Amit Kumar; Singh, Nagendra; Sinha, Mau; Singh, Sanjay; Bhushan, A.; Kaur, Punit; Srinivasan, Alagiri; Sharma, Sujata; Singh, T.P.

doi:10.1074/jbc.m807644200

Cited by 49 publications

(15 citation statements)

References 36 publications

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“…The subsites S1, S2, and S3 are largely similar in both LPO and MPO. The subsite S4 differs slightly, whereas subsite S5 is formed very differently (20), producing markedly different substrate specificities in the two enzymes. Subsite S5 represents the loop Lys-420 -Phe-431, and because S5 is critical in the substrate recognition, the loop Lys-420 -Phe-431 is termed as substrate specific loop.…”

Section: Resultsmentioning

confidence: 99%

“…Among the four mammalian peroxidases, the prosthetic heme group is linked through three covalent bonds in MPO, whereas in LPO, EPO, and thyroid peroxidase only two covalent linkages are formed. So far the detailed crystal structures of only two mammalian peroxidases, MPO and LPO, are known (15)(16)(17)(18)(19)(20). One of the most striking differences between these two mammalian peroxidases is concerned with the basic structural organization in which MPO exists as a covalently linked dimer, whereas LPO is a monomeric protein.…”

Section: Lactoperoxidase (Lpo)mentioning

confidence: 99%

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Binding Modes of Aromatic Ligands to Mammalian Heme Peroxidases with Associated Functional Implications

Singh

Sinha

et al. 2009

Journal of Biological Chemistry

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Lactoperoxidase (LPO) 4 (EC 1.11.1.7) is a member of the family of glycosylated mammalian heme-containing peroxidase enzymes which also includes myeloperoxidase (MPO), eosinophil peroxidase (EPO), and thyroid peroxidase. These enzymes also show functional similarities to non-homologous plant and fungal peroxidases because they follow a similar scheme of reaction (1-3), but their modes of ligand binding differ considerably. Furthermore, the association of the prosthetic heme group in mammalian peroxidases is through covalent bonds (4 -9), whereas covalent linkages are absent in other peroxidases (10 -14). Among the four mammalian peroxidases, the prosthetic heme group is linked through three covalent bonds in MPO, whereas in LPO, EPO, and thyroid peroxidase only two covalent linkages are formed. So far the detailed crystal structures of only two mammalian peroxidases, MPO and LPO,. One of the most striking differences between these two mammalian peroxidases is concerned with the basic structural organization in which MPO exists as a covalently linked dimer, whereas LPO is a monomeric protein.At present the fundamental questions pertaining to mammalian heme peroxidases are (i) what distinguishes between the aromatic ligands that one ligand acts as a substrate, whereas the other ligand works as an inhibitor and (ii) how the substrate and inhibitor specificities differ between two enzymes lactoperoxidase and myeloperoxidase.Lactoperoxidase oxidizes inorganic ions, preferentially thiocyanate (SCN Ϫ ), and to a lesser extent, bromide (Br Ϫ ), whereas in the case of myeloperoxidase the chloride (Cl Ϫ ) ion is a preferred substrate (21,22). The mammalian peroxidases including LPO are also involved in catalyzing the single electron oxidation of various physiologically important organic aromatic substrates including phenols (23, 24), catecholamines, and catechols (25-27) as well as other experimental model compounds such as aromatic amines (28), polychlorinated biphenyls (29), steroid hormones (30 -32), and polycyclic aromatic hydrocarbons (33). However, the mode of binding of aromatic ligands and associated functional implications are not yet clearly understood. Surprisingly, the structural data on the complexes of mammalian peroxidases with aromatic ligands are conspicuously lacking. The only available structural report is on the complex of MPO with salicylhydroxamic acid (SHA) (34). Even in this case, the coordinates of this structure are not available for a detailed analysis. In the case of non-homologous plant peroxidases, a few crystal structures of their complexes with aromatic compounds are available (35-38), but their modes of binding are not very similar to those of mammalian peroxidases because the distal ligand binding sites in mammalian and plant peroxidases differ markedly. In this regard it is pertinent to note

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Section: Resultsmentioning

confidence: 99%

Section: Lactoperoxidase (Lpo)mentioning

confidence: 99%

Binding Modes of Aromatic Ligands to Mammalian Heme Peroxidases with Associated Functional Implications

Singh

Sinha

et al. 2009

Journal of Biological Chemistry

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“…However, the release of free iron does not necessarily require oxidative cleavage of all the three heme-protein covalent bonds, but does require the opening or degradation of heme moiety. HOCl can either attack the covalent tethering points leading to the release of the intact tetrapyrrole ring and/or randomly attack any of the four carbon-methyne bridges [19][20][21][22][23][24]. At the HOCl concentrations used in this study, the intact tetrapyrrole ring was not detected, but instead one tripyrrole, and eleven dipyrroles were identified.…”

Section: Discussionmentioning

confidence: 85%

“…1, B) [78]. Because of these distortions to the porphyrin ring, the α, β, γ, and δ carbon-methyne bridges become more susceptible to HOCl attack, which in turn negatively affects the affinity of the heme iron toward the axial ligands [19][20][21][22][23][24]. The majority of the twelve identified compounds are dipyrrole derivatives formed by HOCl mediated cleavage of the tetrapyrrole ring either through the δ-β axis or through the α-γ axis.…”

Section: Discussionmentioning

confidence: 99%

“…As shown in Fig. 1, the heme prosthetic of MPO is covalently attached to the protein through glutamate (R 1 ), methionine (R 2 ), and aspartate (R 3 ) residues, while LPO and EPO are covalently attached to the protein through glutamate (R 1 ) and aspartate (R 3 ) residues only [19][20][21][22][23][24]. MPO x-ray structure analysis, like EPO and LPO, showed a non-planar configuration of the heme moiety in which pyrrole rings B and D are nearly co-planar, while rings A and C are tilted toward the distal side ( Fig.…”

Section: Introductionmentioning

confidence: 99%

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Disruption of Heme-Peptide Covalent Cross-Linking in Mammalian Peroxidases by Hypochlorous Acid

Maitra

Byun

Andreana

et al. 2012

Free Radical Biology and Medicine

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Dual binding mode of antithyroid drug methimazole to mammalian heme peroxidases – structural determination of the lactoperoxidase–methimazole complex at 1.97 Å resolution

Singh

Sirohi

et al. 2016

FEBS Open Bio

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Lactoperoxidase ( LPO , EC 1.11.1.7 ) is a member of the mammalian heme peroxidase family which also includes thyroid peroxidase ( TPO ). These two enzymes have a sequence homology of 76%. The structure of LPO is known but not that of TPO . In order to determine the mode of binding of antithyroid drugs to thyroid peroxidase, we have determined the crystal structure of LPO complexed with an antithyroid drug, methimazole ( MMZ ) at 1.97 Å resolution. LPO was isolated from caprine colostrum, purified to homogeneity and crystallized with 20% poly(ethylene glycol)‐3350. Crystals of LPO were soaked in a reservoir solution containing MMZ . The structure determination showed the presence of two crystallographically independent molecules in the asymmetric unit. Both molecules contained one molecule of MMZ , but with different orientations. MMZ was held tightly between the heme moiety on one side and the hydrophobic parts of the side chains of Arg255, Glu258, and Leu262 on the opposite side. The back of the cleft contained the side chains of Gln105 and His109 which also interacted with MMZ . In both orientations, MMZ had identical buried areas and formed a similar number of interactions. It appears that the molecules of MMZ can enter the substrate‐binding channel of LPO in two opposite orientations. But once they reach the distal heme pocket, their orientations are frozen due to equally tight packing of MMZ in both orientations. This is a novel example of an inhibitor binding to an enzyme with two orientations at the same site with nearly equal occupancies.

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Structural Evidence of Substrate Specificity in Mammalian Peroxidases

Cited by 49 publications

References 36 publications

Binding Modes of Aromatic Ligands to Mammalian Heme Peroxidases with Associated Functional Implications

Binding Modes of Aromatic Ligands to Mammalian Heme Peroxidases with Associated Functional Implications

Disruption of Heme-Peptide Covalent Cross-Linking in Mammalian Peroxidases by Hypochlorous Acid

Dual binding mode of antithyroid drug methimazole to mammalian heme peroxidases – structural determination of the lactoperoxidase–methimazole complex at 1.97 Å resolution

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