Site‐directed mutants of human myeloperoxidase A topological approach to the heme‐binding site

Jacquet, Alain; Deleersnyder, Virginie; Garcia‐Quintana, Lida; Bollen, Alex; Moguilevsky, Nicole

doi:10.1016/0014-5793(92)80437-l

Cited by 12 publications

(5 citation statements)

References 12 publications

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“…In line with this fact, iron depletion attenuates UPEC pathogenesis in a mouse and rat model of urinary tract infection and PN [44,45]. We found elevated expression of several AMPs with iron sequestering or heme-binding properties, namely calprotectin, NGAL, lactotransferrin, myeloperoxidase and eosinophil peroxidase [46][47][48][49]. Calprotectin is known to be produced and released by neutrophils, monocytes, and macrophages [23] and was found to be upregulated in acute renal allograft failure, AKI [23], obstructive nephropathy (ON) [24], ischemia/reperfusion (I/R) injury [50], staphylococcus infection-associated glomerulonephritis (SAGN) [25], and anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV) [26].…”

Section: Discussionsupporting

confidence: 69%

Proteomic adaptations in the kidney reveal orchestration of local and secreted antimicrobial peptides in human pyelonephritis

Borgards,

Siebels,

Voss

et al. 2023

Preprint

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Pyelonephritis (PN) is a frequent bacterial infection of the kidney and is often associated with severe diseases, organ loss and sepsis. Antibiotics are the cornerstone of therapy, however, increasing antibiotic resistance threatens therapy success and necessitates novel treatment strategies. Various proteins, such as antimicrobial peptides (AMPs), are key molecules of the innate immune response and insights into their regulation may help overcome multi-drug resistance and severe diseases. Using label-free liquid chromatography-tandem mass spectrometry (LC-MS/MS), several cellular, biological, and metabolic processes important for the antimicrobial response were identified, including a significant increase in previously undescribed proteins in human PN with antimicrobial function. Among others, we observed elevation of AMPs, such as calprotectin, azurocidin-1, and cathepsin G in the kidney, which we validated in the urine. Additionally, we observed a negative correlation of azurocidin-1 with plasma levels of C-reactive protein suggesting that the presence in the kidney may protect from severe diseases and systemic inflammation. This study represents the first renal proteomic dataset of human PN, enabling novel insights into the expression of AMPs in the context of PN.

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Section: Discussionsupporting

confidence: 69%

Proteomic adaptations in the kidney reveal orchestration of local and secreted antimicrobial peptides in human pyelonephritis

Borgards,

Siebels,

Voss

et al. 2023

Preprint

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“…The failure to engineer MPO‐like properties into LPO by a single mutation was disappointing inasmuch as the reverse strategy used previously [15, 17], i.e. substituting Met‐409 in MPO by Gln, led to the loss of the chlorinating activity, to a red shift of the Soret peak from 428 nm to 413 nm and to the relative decrease in peroxidase activity.…”

Section: Resultsmentioning

confidence: 99%

“…Previous studies using site‐directed mutagenesis of MPO revealed the nature of the bonds linking the heme group to the amino acid backbone and showed the importance of Met‐409 for enzymatic activity [15–17](Fig. 1 ).…”

Section: Introductionmentioning

confidence: 98%

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Recombinant bovine lactoperoxidase as a tool to study the heme environment in mammalian peroxidases

et al. 1998

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The cDNA encoding bovine lactoperoxidase (LPO) has been expressed in CHO cells. The recombinant LPO was secreted as an enzymatically active single chain molecule presenting two immunoreactive forms of 88 kDa and 82 kDa, differing by their glycosylation. rLPO exhibited the characteristic absorbance spectrum with a Soret peak at 413 nm. Engineering of rLPO into a myeloperoxidase (MPO)-like molecule was attempted by substituting Gln-376 by Met, a residue known to achieve covalent binding with the heme in MPO. However, the resulting bovine LPO mutant failed to acquire the peculiar absorbance spectrum and the chlorinating activity of MPO, underlining the complex nature of interactions in the heme vicinity.z 1998 Federation of European Biochemical Societies.

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“…Site-directed mutagenesis studies [35], which are on the way, will show whether this suggestion is correct.…”

Section: Discussionmentioning

confidence: 96%

Optical spectrum of myeloperoxidase

Floris

Kim

Babcock

et al. 1994

European Journal of Biochemistry

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The optical spectrum of reduced myeloperoxidase (EC 1.11.1.7) displays an unusual red shift of the Soret band which is at 472 nm and the a-band which is at 636 nm. The spectral properties of myeloperoxidase can be modified by means of acid treatment. Upon short exposure to acid (pH 1.7) the red-shifted optical absorption spectrum of the reduced enzyme (A,,, at 472 nm) was blue-shifted (Ama at 448 nm) but the spectrum of the reduced state could be restored by increasing the pH. By contrast, the resonance Raman spectra of both the oxidized and reduced enzyme are essentially the same at both pH 1.7 and pH 7.0. This shows that the optical spectrum and the resonance Raman spectrum are not directly correlated, which we interpret to indicate that the reversible effects of lower pH primarily affect the excited-state energy levels of the macrocycle. The EPR spectrum of the oxidized enzyme showed a reversible conversion from a high-spin rhombic spectrum (8, = 6.7, g, = 5.2) at neutral pH into a more axial high-spin spectrum (gx = g, = 5.8) at low pH.Upon prolonged exposure to acid (20 min) optical absorbance spectra, EPR spectra, resonance Raman spectra and the chlorinating activity were irreversibly affected. We propose that a negatively charged protonatable residue in the proximity of a pyrrole nucleus of the haem group is present that imposes the red shift in the optical absorption spectrum. This is consistent with the available X-ray structure data.Myeloperoxidase is present in large amounts in polymorphonuclear neutrophils and plays an important role in the antimicrobial activity. The enzyme catalyzes the H,O,-dependent peroxidation of chloride to hypochlorous acid, which is a bactericidal agent [l, 21. Myeloperoxidase has also been shown to react very rapidly with peroxynitrite and a new role for the enzyme as a scavenger for peroxynitrite has been proposed [3].The optical absorption spectrum of the reduced enzyme is unique because the Soret band (472nm) and the a-band (636 nm) are red-shifted compared with other haem proteins [4]. The chemical nature of the prosthetic group of myeloperoxidase, which is a haem group, has, however, not been identified, partly due to the fact that the haem group is not readily extractable. This is possibly the result of a covalent linkage between the prosthetic group and the protein [5-71. The crystal structure has been determined at 0.3-nm resolution [S], however, the exact nature of the prosthetic group could not be established from the current X-ray data. It has been suggested that the chromophore is a porphyrin with electronwithdrawing substituents because pyridine haemochrome spectra and the spectra of the enzyme treated with SDS or acid have a strong similarity with that of the haem a spectra of the corresponding form of cytochrome c oxidase [9-131.

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Site‐directed mutants of human myeloperoxidase A topological approach to the heme‐binding site

Cited by 12 publications

References 12 publications

Proteomic adaptations in the kidney reveal orchestration of local and secreted antimicrobial peptides in human pyelonephritis

Proteomic adaptations in the kidney reveal orchestration of local and secreted antimicrobial peptides in human pyelonephritis

Recombinant bovine lactoperoxidase as a tool to study the heme environment in mammalian peroxidases

Optical spectrum of myeloperoxidase

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