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J. Biol. Chem., Vol. 282, Issue 38, 27994-28003, September 21, 2007

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Impact of Two Novel Mutations on the Structure and Function of Human Myeloperoxidase^*

Melissa Goedken, Sally McCormick, Kevin G. Leidal, Kazuo Suzuki, Yosuke Kameoka^¶, Joshua M. Astern^||, Meilan Huang^**, Artem Cherkasov^**, and William M. Nauseef¹

From the Inflammation Program, Department of Medicine, University of Iowa and Veterans Affairs Medical Center, Iowa City, Iowa 52241, National Institute of Infectious Diseases, Toyama 1-23-1, Shinjuku-ku, Tokyo 162 8640, Japan, ^¶Laboratory of Genetic Resources, Division of Biomedical Resources, National Institute of Biomedical Innovation, 7-6-8, Saitoasagi, Ibaraki-City, Osaka, 567-0085, Japan, ^||University of North Carolina, Kidney Center, Chapel Hill, North Carolina 27599, and ^**Division of Infectious Diseases, Department of Medicine, Faculty of Medicine, University of British Columbia, Vancouver V5Z 3J5, British Columbia, Canada

The heme protein myeloperoxidase (MPO) contributes critically to O₂-dependent neutrophil antimicrobial activity. Two Japanese adults were identified with inherited MPO deficiency because of mutations at Arg-499 or Gly-501, conserved residues near the proximal histidine in the heme pocket. Because of the proximity of these residues to a critical histidine in the heme pocket, we examined the biosynthesis, function, and spectral properties of the peroxidase stably expressed in human embryonic kidney cells. Biosynthesis of normal MPO by human embryonic kidney cells faithfully mirrored events previously identified in cells expressing endogenous MPO. Mutant apopro-MPO was 90 kDa and interacted normally with the molecular chaperones ERp57, calreticulin, and calnexin in the endoplasmic reticulum. However, mutant precursors were not proteolytically processed into subunits of MPO, although secretion of the unprocessed precursors occurred normally. Although -[¹⁴C]aminolevulinic acid incorporation demonstrated formation of pro-MPO in both mutants, neither protein was enzymatically active. The Soret band for each mutant was shifted from the normal 430 to 412 nm, confirming that heme was incorporated but suggesting that the number of covalent bonds or other structural aspects of the heme pocket were disrupted by the mutations. These studies demonstrate that despite heme incorporation, mutations in the heme environs compromised the oxidizing potential of MPO.

Received for publication, March 7, 2007 , and in revised form, July 23, 2007.

^* This work was supported by National Institutes of Health Grant HL 53592 and a Merit Review from Veterans Affairs (to W. M. N.). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

¹ To whom correspondence should be addressed: Inflammation Program, and Dept. of Medicine, Roy J. and Lucille A. Carver College of Medicine, University of Iowa, D160 MTF, 2501 Crosspark Road, Coralville, IA 52241. Tel.: 319-335-4278; Fax: 319-335-4194; E-mail: william-nauseef{at}uiowa.edu.

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