tailieunhanh - Báo cáo Y học: Expression of recombinant murine pregnancy-associated plasma protein-A (PAPP-A) and a novel variant (PAPP-Ai) with differential proteolytic activity

Murine pregnancy-associated plasma protein-A (PAPP-A) cDNA encoding a 1545 amino-acid protein has been cloned. We have also identified and cloned cDNA that encodes a novel variant of PAPP-A, PAPP-Ai, carrying a 29-residue highly basic insert. The point of insertion corresponds to a junction between two exons in the human PAPP-A gene. The human intron flanked by these exons does not encode a homologous corresponding insert, which is unique to the mouse. | Eur. J. Biochem. 269 2247-2256 2002 FEBS 2002 doi Expression of recombinant murine pregnancy-associated plasma protein-A PAPP-A and a novel variant PAPP-Ai with differential proteolytic activity Rikke See1 Michael T. Overgaard1 Anni R. Thomsen1 Lisbeth S. Laursen1 Inger M. Olsen1 Lars Sottrup-Jensen1 Jesper Haaning1 Linda C. Giudice2 Cheryl A. Conover3 and Claus Oxvig1 1Department of Molecular and Structural Biology Science Park University of Aarhus Denmark department of Gynecology and Obstetrics Stanford University Stanford CA USA 3Endocrine Research Unit Mayo Clinic and Foundation Rochester USA Murine pregnancy-associated plasma protein-A PAPP-A cDNA encoding a 1545 amino-acid protein has been cloned. We have also identified and cloned cDNA that encodes a novel variant of PAPP-A PAPP-Ai carrying a 29-residue highly basic insert. Tie point of insertion corresponds to a junction between two exons in the human PAPP-A gene. The human intron flanked by these exons does not encode a homologous corresponding insert which is unique to the mouse. The ovrrall eequenre idnntity htlween murine and human PAPP-A is 91 and murine PAPP-A contains sequence motifs previously described in the sequence of human PAPP-A. Through expt esinm m mammalían cells we show that murine PAPP-A and PAPP-Ai are active metalloproteinases both capable of cleaving insulin-like growth factor binding protein IGFBP -4 and -5. Qeavage of IGFBP-4 is dramatically enhanced by the addition of IGF whereas cleavage of IGFBP-5 is slightly inhibited by IGF as previously established with human PAPP-A. Surprisingly however quantitative analyses demonstrate that the murine PAPP-Ai cleaves IGFBP-4 very slowly compared to PAPP-A even though its ability to cleave IGFBP-5 is unaffected by the presence of the insert. By RT-PCR analysis we find that both variants are expressed in several tissues. The level nf mRNA m the mutine paic-enta does not exceed the levels of other tissues .