Identification and structural and functional characterization of human enamelysin (MMP-20).
A cDNA encoding a new human matrix metalloproteinase (MMP) has been cloned from RNA prepared from odontoblastic cells. The open reading frame of the cloned cDNA codes for a polypeptide of 483 amino acids and is extensively similar to the sequence of recently described porcine enamelysin, suggesting that the isolated cDNA codes for the human homologue of this enzyme. Human enamelysin (MMP-20) has a domain organization similar to other MMPs, including a signal peptide, a prodomain with the conserved motif PRCGVPD involved in maintaining enzyme latency, a catalytic domain with a Zn-binding site, and a COOH-terminal fragment similar to the sequence of hemopexin. The calculated molecular mass of human enamelysin is about 54 kDa, which is similar to that of collagenases or stromelysins. However, this human MMP lacks a series of structural features distinctive of these subfamilies of MMPs. The full-length human enamelysin cDNA has been expressed in Escherichia coli, and the purified and refolded recombinant protein is able to degrade synthetic peptides used as substrates of MMPs, confirming that human enamelysin belongs to this family of proteases. Furthermore, the recombinant human enamelysin is able to degrade amelogenin, the major protein component of the enamel matrix. On the basis of its degrading activity on amelogenin, and its highly restricted expression to dental tissues, we suggest that human enamelysin plays a central role in the process of tooth enamel formation. Finally, we have found that the human enamelysin gene (MMP-20) maps to chromosome 11q22, clustered to at least seven other members of the MMP gene family.
Llano
E
E
Departamento de Bioquímica y Biología Molecular, Facultad de Medicina, Universidad de Oviedo, 33006 Oviedo, Spain.
Pendás
A M
AM
Knäuper
V
V
Sorsa
T
T
Salo
T
T
Salido
E
E
Murphy
G
G
Simmer
J P
JP
Bartlett
J D
JD
López-Otín
C
C
D000595:Amino Acid Sequence; D001483:Base Sequence; D002874:Chromosome Mapping; D002880:Chromosomes, Human, Pair 11; D003001:Cloning, Molecular; D018076:DNA, Complementary; D003743:Dental Enamel / Q000737:chemistry; D006801:Humans; D053519:Matrix Metalloproteinase 20; D020782:Matrix Metalloproteinases; D008666:Metalloendopeptidases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid
D018076:DNA, Complementary; D011994:Recombinant Proteins; C507111:MMP20 protein, human; D053519:Matrix Metalloproteinase 20; D020782:Matrix Metalloproteinases; D008666:Metalloendopeptidases