News from an ancient world: two novel astacin metalloproteases from the horseshoe crab.
In this work, we report the cloning, heterologous expression, and characterization of two novel astacin proteases from the chelicerate Limulus polyphemus (horseshoe crab), designated as LAST (Limulus astacin) and LAST_MAM (Limulus astacin containing a MAM domain), respectively. The expression pattern showed ubiquitous occurrence of LAST_MAM, while LAST was predominantly restricted to the eyes and brain, indicating a function in the nervous system. Both enzymes contain the characteristic metzincin-type zinc-binding region and Met turn. While LAST is made up only of the typical prodomain and astacin-like protease domain, LAST_MAM contains an additional MAM (meprin A5 protein tyrosine phosphatase micro) domain, which so far only has been found in few astacins such as the vertebrate meprin Hydra and squid enzymes, and in a number of other extracellular proteins such as A5 protein and tyrosine phosphatase micro. These gave rise to the designation MAM for this protein module. MAM domains have been shown to be responsible for protein oligomerization in meprin proteases and tyrosine phosphatase micro. Since the horseshoe crab has kept its body plan for almost half a billion years, it is therefore a privileged organism for the study of protease evolution. In this context, we could show by phylogenetic analysis that this protease is not related to the other MAM-domain-containing astacins indicating different evolutionary origins of these proteins. Moreover, we clearly demonstrated the divergent evolvement of the MAM module itself, and not only with regard to proteases. However, there are some unique functional features that are not shared by other members of this protein family. For example, LAST_MAM is the only astacin protease known so far that is active in its zymogen form, indicating that the presence of the N-terminal propeptide does not prevent proteolytic activity.
Becker-Pauly
Christoph
C
Institute of Zoology, Cell and Matrix Biology, Johannes Gutenberg-University, Mainz, Germany. beckerpa@uni-mainz.de
Bruns
Bernd Cem
BC
Damm
Olga
O
Schütte
André
A
Hammouti
Kada
K
Burmester
Thorsten
T
Stöcker
Walter
W
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002364:Caseins / Q000378:metabolism; D002460:Cell Line; D003001:Cloning, Molecular; D024042:Collagen Type I / Q000378:metabolism; D018076:DNA, Complementary / Q000235:genetics; D019143:Evolution, Molecular; D016326:Extracellular Matrix Proteins / Q000378:metabolism; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D006737:Horseshoe Crabs / Q000201:enzymology*; D006877:Hydroxamic Acids / Q000494:pharmacology; D007313:Insecta / Q000166:cytology; D008666:Metalloendopeptidases / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009420:Nervous System / Q000201:enzymology; D009842:Oligopeptides / Q000494:pharmacology; D010802:Phylogeny; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D017434:Protein Structure, Tertiary; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D040681:Structural Homology, Protein
D002364:Caseins; D024042:Collagen Type I; D018076:DNA, Complementary; D016326:Extracellular Matrix Proteins; D006877:Hydroxamic Acids; D009842:Oligopeptides; D012333:RNA, Messenger; C026018:azocasein; C090221:prolyl-leucyl-glycine hydroxamate; D008666:Metalloendopeptidases; C071288:astacin; C010531:actinonin