Cloning and deduced amino acid sequence of a novel cartilage protein (CILP) identifies a proform including a nucleotide pyrophosphohydrolase.
The cDNA cloning and expression in vitro and in eukaryotic cells of a novel protein isolated from human articular cartilage, cartilage intermediate layer protein (CILP) is described. A single 4. 2-kilobase mRNA detected in human articular cartilage encodes a polypeptide of 1184 amino acids with a calculated molecular mass of 132.5 kDa. The protein has a putative signal peptide of 21 amino acids, and is a proform of two polypeptides. The amino-terminal half corresponds to CILP (molecular mass of 78.5 kDa, not including post-translational modifications) and the carboxyl-terminal half corresponds to a protein homologous to a porcine nucleotide pyrophosphohydrolase, NTPPHase (molecular mass of 51.8 kDa, not including post-translational modifications). CILP has 30 cysteines and six putative N-glycosylation sites. The human homolog of porcine NTPPHase described here contains 10 cysteine residues and two putative N-glycosylation sites. In the precursor protein the NTPPHase region is immediately preceded by a tetrapeptide conforming to a furin proteinase cleavage consensus sequence. Expression of the full-length cDNA in a cell-free translation system and in COS-7 or EBNA cells indicates that the precursor protein is synthesized as a single polypeptide chain that is processed, possibly by a furin-like protease, into two polypeptides upon or preceding secretion.
Lorenzo
P
P
Department of Cell and Molecular Biology, Section for Connective Tissue Biology, University of Lund, P.O.Box 94, S-221 00 Lund, Sweden.
Neame
P
P
Sommarin
Y
Y
Heinegård
D
D
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D019556:COS Cells; D002356:Cartilage / Q000737:chemistry*; D019902:Chondrocytes / Q000378:metabolism; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D016326:Extracellular Matrix Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D006023:Glycoproteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D008969:Molecular Sequence Data; D010449:Peptide Mapping; D016133:Polymerase Chain Reaction; D011498:Protein Precursors / Q000235:genetics*; D011499:Protein Processing, Post-Translational; D011755:Pyrophosphatases / Q000235:genetics*; D011994:Recombinant Proteins / Q000096:biosynthesis; D015183:Restriction Mapping; D017421:Sequence Analysis; D017386:Sequence Homology, Amino Acid
D018076:DNA, Complementary; D016326:Extracellular Matrix Proteins; D006023:Glycoproteins; D011498:Protein Precursors; D011994:Recombinant Proteins; C114607:CILP protein, human; D011755:Pyrophosphatases; C021899:nucleotide pyrophosphatase