title: Site specificity of DSP-PP cleavage by BMP1.
abstract: Bone morphogenic protein 1 (BMP1), a metalloproteinase, is known to cleave a wide variety of extracellular matrix proteins, suggesting that a consensus substrate cleavage amino acid sequence might exist. However, while such a consensus sequence has been proposed based on P4 to P4' (i.e. the four amino acids flanking either side of the BMP1 cleavage site; P4P3P2P1|P1'P2'P3'P4') sequence homologies between two BMP1 substrates, dentin matrix protein 1 and dentin sialoprotein phosphophoryn (DSP-PP) (i.e. xMQx|DDP), no direct testing has so far been attempted. Using an Sf9 cell expression system, we have been able to produce large amounts of uncleaved DSP-PP. Point mutations introduced into this recombinant DSP-PP were then tested for their effects on DSP-PP cleavage by either Sf9 endogenous tolloid-related protein 1 (TLR-1) or by its human homolog, BMP1. Here, we have measured DSP-PP cleavage efficiencies after modifications based on P4-P4' sequence comparisons with dentin matrix protein 1, as well as for prolysyl oxidase and chordin, two other BMP1 substrates. Our results demonstrate that any mutations within or outside of the DSP-PP P4 to P4' cleavage site can block, impair or accelerate DSP-PP cleavage, and suggest that its BMP1 cleavage site is highly conserved in order to regulate its cleavage efficiency, possibly with additional assistance from its conserved exosites. Thus, BMP1 cleavage cannot be based on a consensus substrate cleavage site.
authors.affiliation: Department of Cariology, Restorative Sciences and Endodontics, School of Dentistry, University of Michigan , Ann Arbor, MI , USA and.
authors.forename: Robert T
authors.initials: RT
authors.lastname: Yang
authors.forename: Glendale L
authors.initials: GL
authors.lastname: Lim
authors.forename: Colin T
authors.initials: CT
authors.lastname: Yee
authors.forename: Robert S
authors.initials: RS
authors.lastname: Fuller
authors.forename: Helena H
authors.initials: HH
authors.lastname: Ritchie
mesh_terms: D000595:Amino Acid Sequence; D055395:Bone Morphogenetic Protein 1 / Q000378:metabolism*; D016326:Extracellular Matrix Proteins / Q000737:chemistry / Q000378:metabolism*; D006023:Glycoproteins / Q000378:metabolism; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins / Q000378:metabolism; D010750:Phosphoproteins / Q000737:chemistry / Q000378:metabolism*; D011498:Protein Precursors / Q000378:metabolism; D012795:Sialoglycoproteins / Q000737:chemistry / Q000378:metabolism*; D000099311:Dentin Sialophosphoprotein
chemical_list: D055395:Bone Morphogenetic Protein 1; D016326:Extracellular Matrix Proteins; D006023:Glycoproteins; D036341:Intercellular Signaling Peptides and Proteins; D010750:Phosphoproteins; D011498:Protein Precursors; D012795:Sialoglycoproteins; D000099311:Dentin Sialophosphoprotein; C017630:phosphophoryn; C054234:chordin; C525705:BMP1 protein, human
keywords: BMP1; DMP1; chordin; dentin sialophosphoprotein; prolysyl oxidase