Myeloperoxidase, Recombinant, Human (MPO, pANCA)

Myeloperoxidase (MPO) is a heme-containing enzyme belonging to the XPO subfamily of peroxidases. It is an abundant neutrophil and monocyte glycoprotein that catalyzes the hydrogen peroxide-dependent conversion of chloride, bromide, and iodide to multiple reactive species (1). Post-translational processing of MPO involves the insertion of a heme moiety and the proteolytic removal of both a propeptide and a 6 aa internal peptide (2). This results in a disulfide-linked dimer composed of a 60 kD heavy and 12kD light chain that associate into a 150 kD enzymatically active tetramer. The tetramer contains two heme groups and one disulfide bond between the heavy chains (2). Alternate splicing generates two additional isoforms of MPO, one with a 32 aa insertion in the light chain, and another with a deletion of the signal sequence and part of the propeptide (3). Human and mouse MPO share 87% aa sequence identity. MPO activity results in protein nitrosylation and the formation of 3-chlorotyrosine and dityrosine crosslinks (4-6). Modification of ApoB100, as well as the lipid and cholesterol components of LDL and HDL, promotes the development of atherosclerosis (5, 7-9). MPO is also associated with a variety of other diseases (1), and inhibits vasodilation in inflammation by depleting the levels of NO (10). Serum albumin functions as a carrier protein during MPO movement to the basolateral side of epithelial cells (11). MPO is stored in neutrophil azurophilic granules. Upon cellular activation, it is deposited into pathogen-containing phagosomes (2). While mice lacking MPO are impaired in clearing select microbial infections, MPO deficiency in humans does not necessarily result in heightened susceptibility to infections (12, 13). Source: Human CD33 Signal Peptide (Met 1-Ala 16), Human Myeloperoxidase (Ala 49-Ser 745), HHHHHHHHHH, A DNA sequence encoding the signal peptide from human CD33 joined with the human Myeloperoxidase (Ala 49-Ser 745, Accession # P05164) was fused with a 10x histidine tag at the C-terminus. The chimeric protein was expressed in a mouse myeloma cell line, NS0. , Molecular Mass: , The recombinant human Myeloperoxidase starts at Ala 49 and has a calculated molecular mass of approximately 80kD. As a result of glycosylation, the protein migrates as an ~80-90 kD protein in SDS-PAGE under reducing conditions. In gel filtration chromatography, the recombinant protein migrates as a tetramer., Endotoxin Level: < 1.0 EU per 1ug of the protein as determined by the LAL method. Activity: Measured by its ability to oxidize guaiacol in the presence of hydrogen peroxide (Desser, R.K., et al. 1972 Arch.Biochem. Biophys. 148: 452-65). See Activity Assay Protocol on next page for details. The specific activity of the recombinant human Myeloperoxidase is > 5000 pmoles/min/ug as defined in the assay. , Storage and Stability: Lyophilized powder may be stored at -20/-70°C. Reconstitute to nominal volume by adding ddH20 and 0.1% BSA or HSA . Aliquot and store at -20/-70°C. Reconstituted product is stable for 3 months at -20/-70°C. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap. Further dilutions can be made in assay buffer.