Identification and characterization of human archaemetzincin-1 and -2, two novel members of a family of metalloproteases widely distributed in Archaea.
Systematic analysis of degradomes, the complete protease repertoires of organisms, has demonstrated the large and growing complexity of proteolytic systems operating in all cells and tissues. We report here the identification of two new human metalloproteases that have been called archaemetzincin-1 (AMZ1) and archaemetzincin-2 (AMZ2) to emphasize their close relationship to putative proteases predicted by bioinformatic analysis of archaeal genomes. Both human proteins contain a catalytic domain with a core motif (HEXXHXXGX3CX4CXMX17CXXC) that includes an archetypal zinc-binding site, the methionine residue characteristic of metzincins, and four conserved cysteine residues that are not present at the equivalent positions of other human metalloproteases. Analysis of genome sequence databases revealed that AMZs are widely distributed in Archaea and vertebrates and contribute to the defining of a new metalloprotease family that has been called archaemetzincin. However, AMZ-like sequences are absent in a number of model organisms from bacteria to nematodes. Phylogenetic analysis showed that these enzymes have undergone a complex evolutionary process involving a series of lateral gene transfer, gene loss, and genetic duplication events that have shaped this novel family of metalloproteases. Northern blot analysis showed that AMZ1 and AMZ2 exhibit distinct expression patterns in human tissues. AMZ1 is mainly detected in liver and heart whereas AMZ2 is predominantly expressed in testis and heart, although both are also detectable at lower levels in other tissues. Both human enzymes were produced in Escherichia coli, and the purified recombinant proteins hydrolyzed synthetic substrates and bioactive peptides, demonstrating that they are functional proteases. Finally, these activities were abolished by inhibitors of metalloproteases, providing further evidence that AMZs belong to this catalytic class of proteolytic enzymes.
Departamento de Bioquímica y Biología Molecular, Facultad de Medicina, Instituto Universitario de Oncología, Universidad de Oviedo, 33006 Oviedo, Spain.
Araceli
A
Díaz-Perales
Víctor
V
Quesada
Juan R
JR
Peinado
Alejandro P
AP
Ugalde
Jesús
J
Alvarez
María F
MF
Suárez
F Xavier
FX
Gomis-Rüth
Carlos
C
López-Otín
D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D001105:Archaea; D001483:Base Sequence; D015152:Blotting, Northern; D002384:Catalysis; D020134:Catalytic Domain; D019295:Computational Biology; D018076:DNA, Complementary / Q000378:metabolism; D019143:Evolution, Molecular; D018507:Gene Expression Regulation, Developmental; D020745:Genome, Archaeal; D006801:Humans; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D008099:Liver / Q000378:metabolism; D008297:Male; D013058:Mass Spectrometry; D045726:Metalloproteases / Q000737:chemistry* / Q000502:physiology; D008969:Molecular Sequence Data; D009206:Myocardium / Q000378:metabolism; D010455:Peptides / Q000737:chemistry; D010802:Phylogeny; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D013737:Testis / Q000378:metabolism; D014018:Tissue Distribution; D014357:Trypsin / Q000494:pharmacology
D018076:DNA, Complementary; D010455:Peptides; D011994:Recombinant Proteins; C504248:AMZ1 protein, human; C504249:AMZ2 protein, human; D045726:Metalloproteases; D014357:Trypsin