Discovery and characterization of human antibody inhibitors of pregnancy-associated plasma protein-A
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Ting Chen
Abstract
Pregnancy-associated plasma protein-A (PAPP-A) is a metalloprotease that cleaves insulin-like growth factor-binding proteins (IGFBPs) to release bioactive levels of free insulin-like growth factor. Specific and potent inhibitors of PAPP-A may further elucidate the biological functions of this protease and could prove to be of therapeutic value. Phage display was used to discover fully human antibody inhibitors of PAPP-A activity towards IGFBP4 cleavage. Estimates of the inhibition constants for these antibodies were subsequently determined using a novel continuous assay of PAPP-A protease activity that uses an internally quenched synthetic peptide substrate (DX-1655). DX-1655 was hydrolyzed by PAPP-A with a Km of 33 μM and a kcat of 0.3 s-1 (kcat/Km=9.1×103 M-1 s-1). PAPP-A activity towards DX-1655 displays a bell-shaped pH profile, with pKa values of 8.2 and 10.8 and a maximum rate at approximately pH 9.5. Using this continuous assay, we measured apparent Ki values of 1.7±0.2 and 7.4±1.5 nM for the F2 and D9 antibodies, respectively.
©2007 by Walter de Gruyter Berlin New York
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Articles in the same Issue
- Mid-region parathyroid hormone-related protein (PTHrP) and gene expression of MDA-MB231 breast cancer cells
- Characterization of peptidyl-tRNA hydrolase encoded by open reading frame Rv1014c of Mycobacterium tuberculosis H37Rv
- Calcium signalling in the regulation of PGC-1α, PDK4 and HKII mRNA expression
- Expression of the protein phosphatase 1 inhibitor KEPI is downregulated in breast cancer cell lines and tissues and involved in the regulation of the tumor suppressor EGR1 via the MEK-ERK pathway
- Nitric oxide modulates expression of extracellular matrix genes linked to fibrosis in kidney mesangial cells
- Discovery and characterization of human antibody inhibitors of pregnancy-associated plasma protein-A
- The protease domain of procollagen C-proteinase (BMP1) lacks substrate selectivity, which is conferred by non-proteolytic domains
- Two α subunits and one β subunit of meprin zinc-endopeptidases are differentially expressed in the zebrafish Danio rerio
- Increase in kinins on post-exercise hypotension in normotensive and hypertensive volunteers
- Inhibition of cathepsin L-like proteases by cathepsin V propeptide
- Clinical chemistry reference database for Wistar rats and C57/BL6 mice
