title: Expression and purification of recombinant protein related to DAN and cerberus (PRDC).
abstract: Bone morphogenetic proteins (BMPs) are secreted protein ligands that control numerous biological processes, such as cell differentiation and cell proliferation. Ligands are regulated by a large number of structurally diverse extracellular antagonists. PRDC or protein related to DAN and cerberus is a BMP antagonist of the DAN family, which is defined by a conserved pattern of cysteine residues that form a ring structure. Here we present the expression and purification of recombinant mouse PRDC (mPRDC) from bacterial (Escherichia coli) inclusion bodies through oxidative refolding. Functional mPRDC was isolated from a nonfunctional component through reverse phase chromatography and shown to inhibit BMP2 and BMP4 in a cell-based luciferase reporter assay. Recombinant mPRDC also bound directly to BMP2, BMP4 and BMP7, but not activin A. Furthermore, circular dichroism indicated that mPRDC is folded and contains a higher than anticipated helical content for a DAN family member protein.
authors.affiliation: Department of Molecular Genetics, Biochemistry and Microbiology, University of Cincinnati Medical Sciences Building, Cincinnati, OH 45267, United States.
authors.forename: Chandramohan
authors.initials: C
authors.lastname: Kattamuri
authors.forename: David M
authors.initials: DM
authors.lastname: Luedeke
authors.forename: Thomas B
authors.initials: TB
authors.lastname: Thompson
mesh_terms: D000818:Animals; D019485:Bone Morphogenetic Proteins / Q000037:antagonists & inhibitors / Q000502:physiology; D002460:Cell Line; D056148:Chromatography, Reverse-Phase; D016207:Cytokines; D015870:Gene Expression; D017930:Genes, Reporter; D057927:Hydrophobic and Hydrophilic Interactions; D002479:Inclusion Bodies / Q000737:chemistry; D008156:Luciferases / Q000096:biosynthesis / Q000235:genetics; D051379:Mice; D011485:Protein Binding; D058849:Protein Refolding; D017433:Protein Structure, Secondary; D011506:Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism* / Q000494:pharmacology; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism / Q000494:pharmacology; D015398:Signal Transduction; D016212:Transforming Growth Factor beta / Q000737:chemistry
chemical_list: D019485:Bone Morphogenetic Proteins; D016207:Cytokines; C490088:Grem2 protein, mouse; D011506:Proteins; D011994:Recombinant Proteins; D016212:Transforming Growth Factor beta; D008156:Luciferases