Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.4.24.11 (CD10)
 9,792 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The phosphate-regulating gene with homologies to endopeptidases on the X chromosome (PHEX) is a member of the neutral endopeptidase family, which is expressed predominantly on the plasma membranes of mature osteoblasts and osteocytes. Although it is known that the loss of PHEX function results in X-linked hypophosphatemic rickets, characterized by abnormal bone matrix mineralization and renal phosphate wasting, little is known about how PHEX is regulated. We therefore sought to determine whether the murine PHEX gene is regulated by glucocorticoids (GCs), which are known to influence phosphate homeostasis and bone metabolism. Northern blot analysis revealed increased PHEX mRNA expression in GC-treated suckling mice (1.5-fold) and in rat osteogenic sarcoma (UMR-106) cells (2.5-fold). An increase was also seen in PHEX promoter activity in transiently transfected UMR-106 cells with GC treatment. Analysis of nested promoter deletions revealed that an atypical GC response element was located between -337 and -315 bp. Mutational analysis and electrophoretic mobility shift assays further identified -326 to -321 bp as a site involved in GC regulation. Supershift analyses and electrophoretic mobility shift assay competition studies indicated that the core binding factor alpha1-subunit transcription factor is able to bind to this region and may therefore play a role in the GC response of the murine PHEX gene.
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PMID:Glucocorticoid regulation of the murine PHEX gene. 1211 May 21

An efficient production method of heme-iron-enriched peptide was developed based on enzymatic hydrolysis. Hemoglobin hydrolysis, carried out stepwise with commercially available exopeptidase and endopeptidase, resulted in an increased degree of hydrolysis (DH). Exopeptidase-catalyzed protein hydrolysis formed low molecular weight peptides and amino acids. Different process parameters including dialysis and ultra- and diafiltration were evaluated. Heme/peptide ratio increased as molecular weight cut-off (MWCO) of the dialysis membrane increased. When the hydrolysate was dialyzed against sodium phosphate buffer, a higher heme/ peptide ratio was obtained. The heme/peptide ratio of the hydrolysate reached up to 25.4% when the dialysis was carried out with a membrane of 12-14 kDa MWCO. Also, the ratio was improved by the use of ultrafiltration and diafiltration on the pilot-scale.
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PMID:Process development for heme-enriched peptide by enzymatic hydrolysis of hemoglobin. 1213 70

We investigated whether the absence of Phex (phosphate-regulating gene with homologies to endopeptidases on the X chromosome) in the Hyp mouse affects the expression and activity of neprilysin (NEP) and of endothelin-converting enzyme-like endopeptidase (ECEL1/DINE) in bone marrow stromal cells (BMSC) and osteoblasts (Ob). Total NEP-like activity was higher in Ob than in BMSC regardless of genotype, and Hyp cells showed higher activities than normal. Conditioned media (CM) from Hyp BMSC and Ob inhibited inorganic phosphate (P(i)) uptake by mouse proximal tubule cells, and incubating Hyp Ob with phosphoramidon prevented the production of the inhibitor of renal P(i) uptake. A linear relationship was observed between the NEP-like activity of Hyp and normal cells and the inhibition of P(i) uptake. NEP and ECEL1/DINE mRNA levels were higher in Hyp cells than in normal cells, and in situ hybridization of ECEL1/DINE confirmed higher levels of expression in the Hyp mouse than in normal cells. In conclusion, we observed a correlation between the inhibition of P(i) uptake by CM from Hyp cells and elevated NEP-like activities.
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PMID:Role of abnormal neutral endopeptidase-like activities in Hyp mouse bone cells in renal phosphate transport. 1237 2

The PHEX gene (phosphate-regulating gene with homologies to endopeptidases on the X chromosome) encodes a protein (PHEX) with structural homologies to members of the M13 family of zinc metallo-endopeptidases. Mutations in the PHEX gene are responsible for X-linked hypophosphataemia in humans. However, the mechanism by which loss of PHEX function results in the disease phenotype, and the endogenous PHEX substrate(s) remain unknown. In order to study PHEX substrate specificity, combinatorial fluorescent-quenched peptide libraries containing o -aminobenzoic acid (Abz) and 2,4-dinitrophenyl (Dnp) as the donor-acceptor pair were synthesized and tested as PHEX substrates. PHEX showed a strict requirement for acidic amino acid residues (aspartate or glutamate) in S(1)' subsite, with a strong preference for aspartate. Subsites S(2)', S(1) and S(2) exhibited less defined specificity requirements, but the presence of leucine, proline or glycine in P(2)', or valine, isoleucine or histidine in P(1) precluded hydrolysis of the substrate by the enzyme. The peptide Abz-GFSDYK(Dnp)-OH, which contains the most favourable residues in the P(2) to P(2)' positions, was hydrolysed by PHEX at the N-terminus of aspartate with a k(cat)/ K(m) of 167 mM(-1) x s(-1). In addition, using quenched fluorescence peptides derived from fibroblast growth factor-23 and matrix extracellular phosphoglycoprotein sequences flanked by Abz and N -(2,4-dinitrophenyl)ethylenediamine, we showed that these physiologically relevant proteins are potential PHEX substrates. Finally, our results clearly indicate that PHEX does not have neprilysin-like substrate specificity.
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PMID:Human recombinant endopeptidase PHEX has a strict S1' specificity for acidic residues and cleaves peptides derived from fibroblast growth factor-23 and matrix extracellular phosphoglycoprotein. 1267 20

We present a new antibody-directed enzyme prodrug therapy strategy (ADEPT) based on a post-proline cleaving endopeptidase and prodrugs, in which cytotoxic moieties are linked to a proline-containing peptide. Human prolyl endopeptidase was expressed in Escherichia coli and purified to homogeneity. The enzyme was active in buffer and in human serum but was rapidly thermally inactivated by incubation at 37 degrees C, thus preventing applications in vivo. While prolyl endopeptidase display on filamentous phage abolished viral infectivity and prevented directed evolution strategies based on phage display, we robotically screened 10752 individual colonies of mutant enzymes using a fluorogenic assay to improve enzyme stability. A single amino acid mutation (Glu289 --> Gly) improved protein stability, resulting in a half-life of 16 h at 37 degrees C in phosphate buffer. Two prodrugs were synthesized, in which an N-protected glycine-proline dipeptide was covalently coupled to doxorubicin and melphalan. (Benzyloxycarbonyl)glycylprolylmelphalan, but not the more sterically hindered doxorubicin prodrug, could be efficiently activated by prolyl endopeptidase [specific activity = 813.3 nmol min(-1) (mg of enzyme)(-1) at 25 degrees C]. The melphalan prodrug was essentially nontoxic to CHO, F9 teratocarcinoma, MCF7 breast adenocarcinoma, and p3U1 mouse myeloma cells up to millimolar concentrations, while prodrug incubation with the engineered prolyl endopeptidase mutant led to a cell killing profile superimposable to the one of melphalan. The prolyl endopeptidase mutant was then chemically coupled to the human antibody L19, specific to the EDB domain of fibronectin, a marker of angiogenesis. The resulting immunoconjugate retains antigen binding and enzymatic activity, thus opening the way to anticancer ADEPT applications.
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PMID:Engineering a thermostable human prolyl endopeptidase for antibody-directed enzyme prodrug therapy. 1514 13

Inorganic phosphate (Pi) is required for cellular function and skeletal mineralization. Serum Pi level is maintained within a narrow range through a complex interplay between intestinal absorption, exchange with intracellular and bone storage pools, and renal tubular reabsorption. The crucial regulated step in Pi homeostasis is the transport of Pi across the renal proximal tubule. Type II sodium-dependent phosphate (Na/Pi) cotransporter (NPT2) is the major molecule in the renal proximal tubule and is regulated by Pi, parathyroid hormone and by 1,25-dihydroxyvitamin D. Recent studies of inherited and acquired hypophosphatemia [X-linked hypophosphatemic rickets/osteomalacia (XLH), autosomal dominant hypophosphatemic rickets/osteomalacia (ADHR) and tumor-induced rickets/osteomalacia (TIO)], which exhibit similar biochemical and clinical features, have led to the identification of novel genes, PHEX and FGF23, that play a role in the regulation of Pi homeostasis. The PHEX gene, which is mutated in XLH, encodes an endopeptidase, predominantly expressed in bone and teeth, but not in kidney. FGF-23 may be a substrate of this endopeptidase and may therefore accumulate in patients with XLH. In the case of ADHR mutations in the furin cleavage site, which prevent the processing of FGF-23 into fragments, lead to the accumulation of a "stable" circulating form of the peptide which also inhibits renal Pi reabsorption. In the case of TIO, ectopic overproduction of FGF-23 overwhelms its processing and degradation by PHEX, leading to the accumulation of FGF-23 in the circulation and inhibition of renal Pi reabsorption. Mice homozygous for severely hypomorphic alleles of the Klotho gene exhibit a syndrome resembling human aging, including atherosclerosis, osteoporosis, emphysema, and infertility. The KLOTHO locus is associated with human survival, defined as postnatal life expectancy, and longevity, defined as life expectancy after 75. In considering the relationship of klotho expression to the dietary Pi level, the klotho protein seemed to be negatively controlled by dietary Pi.
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PMID:Inorganic phosphate homeostasis and the role of dietary phosphorus. 1525 67

X-linked hypophosphatemia (XLH) and autosomal dominant hypophosphatemic rickets (ADHR) are characterized by renal phosphate wasting, rickets, and osteomalacia. ADHR is caused by gain of function mutations in the fibroblast growth factor 23 gene (FGF23). During secretion, FGF23 is processed at the C-terminus between amino acids 179 and 180. The cleavage site is mutated in ADHR, preventing processing of FGF23. Here, we show that FGF23 is likely to be cleaved by subtilisin-like proprotein convertases (SPC) as cleavage can be inhibited by a specific SPC inhibitor in HEK293 cells. SPCs, which are widely expressed, were demonstrated to be also present in HEK293 cells as well as in osteoblasts. XLH is caused by loss of function mutations in the putative endopeptidase PHEX. It was tempting to speculate that FGF23 is a substrate of PHEX, but studies have been inconclusive so far. Here, we used a secreted form of PHEX (secPHEX) and tagged and untagged FGF23 constructs for co-incubation experiments. These experiments provided evidence against cleavage of intact FGF23(25-251) as well as of N-terminal (FGF23(25-179)) and C-terminal (FGF23(180-251)) fragments by the endopeptidase PHEX.
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PMID:FGF23 is processed by proprotein convertases but not by PHEX. 1526 97

The PHEX gene encodes an endopeptidase expressed in osteoblasts that inactivates an uncharacterized peptide hormone, phosphatonin, which suppresses bone mineralization as well as renal phosphate reabsorption and vitamin D bioactivation. We demonstrate that 1alpha-25-dihydroxyvitamin D (1,25(OH)2D3), the, active renal vitamin D metabolite, decreases PHEX mRNA in the rat osteoblastic cell line, UMR-106, as well as in mouse calvaria. Promoter/reporter construct analysis of the murine PHEX gene in transfected UMR-106 cells localized the repressive effect of 1,25(OH)2D3 to the -133 to -74 bp region, and gel mobility shift experiments revealed that 1,25(OH)2D3 treatment of the cells diminished the binding of a nuclear protein(s) to a stretch of 17 adenines from bp -116 to -100 in the proximal PHEX promoter. Either overexpression of a dominant-negative vitamin D receptor (VDR) or deletion of this sequence of 17 A-T base pairs abolished the repressive effect of 1,25(OH)2D3 by attenuating basal promoter activity, indicating that this region mediates the 1,25(OH)2D3 response and is involved in basal transcription. South-western blot analysis and DNA affinity purification show that an unidentified 110 kDa nuclear protein binds to the poly(A) element. Because 1,25(OH)2D3-liganded VDR neither binds to the polyadenine region of the PHEX promoter nor directly influences the association of the 110 kDa transfactor, we conclude that 1,25(OH)2D3 indirectly decreases PHEX expression via VDR-mediated repression (or modification) of this novel transactivator. Thus, we have identified a cis-element required for PHEX gene transcription that participates in negative feedback control of PHEX expression and thereby modulates the actions of phosphatonin.
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PMID:1,25-dihydroxyvitamin D3 down-regulation of PHEX gene expression is mediated by apparent repression of a 110 kDa transfactor that binds to a polyadenine element in the promoter. 1533 62

Inorganic phosphate (Pi) is required for cellular function and skeletal mineralization. Serum Pi level is maintained within a narrow range through a complex interplay between intestinal absorption, exchange with intracellular and bone storage pools, and renal tubular reabsorption. Pi is abundant in the diet, and intestinal absorption of Pi is efficient and minimally regulated. The kidney is a major regulator of Pi homeostasis and can increase or decrease its Pi reabsorptive capacity to accommodate Pi need. The crucial regulated step in Pi homeostasis is the transport of Pi across the renal proximal tubule. Type II sodium-dependent phosphate (Na/Pi) cotransporter (NPT2) is the major molecule in the renal proximal tubule and is regulated by hormones and nonhormonal factors. Recent studies of inherited and acquired hypophosphatemia which exhibit similar biochemical and clinical features, have led to the identification of novel genes, phosphate regulating gene with homologies to endopeptidases on the X chromosome (PHEX) and fibroblast growth factor-23 (FGF-23), that play a role in the regulation of Pi homeostasis. The PHEX gene encodes an endopeptidase, predominantly expressed in bone and teeth but not in kidney. FGF-23 may be a substrate of this endopeptidase and inhibit renal Pi reabsorption. In a survey in the United States and in Japan, the amount of phosphorus from food is gradually increasing. It is thought that excess amounts of phosphorus intake for long periods are a strong factor in bone impairment and ageing. The restriction of phosphorus intake seems to be important under low calcium intake to keep QOL on high level.
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PMID:The regulation and function of phosphate in the human body. 1563 Feb 24

A case of an adult patient with vitamin D-resistant osteomalacia or X-linked hypophosphatemic osteomalacia (XLH) with diffuse calcification of entheses is reported. XLH is the most frequent cause of rickets in developed countries. It is characterized by an impaired renal transport of the phosphate and mutation of PFEX (phosphate regulating gene, with homologies to endopeptidase on the X-chromosome). In childhood, the classic clinical presentation includes short stature and bow leg. While at this age the main radiographic features are characterised by rickets, in adult life they are dominated by a generalised calcific enthesopathy. Concerning the pathogenesis of the enthesopathic lesions of XLH, no convincing hypothesis has yet been made. As in our patient, the extension and the severity of enthesopathy seems not related to the severity of the biochemical changes nor to the treatment with calcitriol. The calcified enthesopathy is an integral part of XLH and it is possible that it is found in adult because many years are necessary to produce it.
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PMID:[The enthesopathy of vitamin D-resistant osteomalacia in adults]. 1577 47


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