Differences in the activation mechanism between the alpha and beta subunits of human meprin.
Meprins are zinc-endopeptidases of the astacin family, which are expressed as membrane-bound or secreted forms in renal and intestinal brush-border membranes of mouse, rat and man. There are two types of meprin subunits, alpha and beta, which form disulfide-bonded homo- and heterodimers; further oligomerization is mediated by non-covalent interactions. Both subunits are translated as proenzymes that have to be activated by removal of an N-terminal propeptide. In the gut, the most probable activator is trypsin. In addition, plasmin has been shown to activate the human alpha subunit in colorectal cancer tissue. In the present study we have overexpressed the human meprin alpha subunit and a His-tagged soluble tail-switch-mutant of meprin beta in Baculovirus-infected insect cells. The recombinant homo-oligomeric proteins were purified by gel filtration and affinity chromatography with yields of up to 10 mg/l cell culture medium and analyzed with regard to their activation mechanism. While both alpha and beta homo-oligomers are activated by trypsin, only meprin alpha homo-oligomers are processed to their mature form by plasmin. These results indicate a different accessibility of the propeptide in meprin homo-oligomers and suggest an explanation for the appearance of meprin hetero-oligomers consisting of active alpha, but latent beta subunits.
Becker
Christoph
C
Institut für Zoophysiologie, Molekulare Physiologie, Westfälische Wilhelms-Universität Münster, Hindenburgplatz 55, D-48143 Münster, Germany.
Kruse
Markus-N
MN
Slotty
Kristina A
KA
Köhler
Danny
D
Harris
J Robin
JR
Rösmann
Sandra
S
Sterchi
Erwin E
EE
Stöcker
Walter
W
D002850:Chromatography, Gel; D004789:Enzyme Activation / Q000187:drug effects; D004792:Enzyme Precursors / Q000378:metabolism; D005341:Fibrinolysin / Q000494:pharmacology; D006801:Humans; D015151:Immunoblotting; D008666:Metalloendopeptidases / Q000737:chemistry / Q000378:metabolism*; D008854:Microscopy, Electron; D021122:Protein Subunits / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D014357:Trypsin / Q000494:pharmacology; D015032:Zinc / Q000737:chemistry / Q000378:metabolism*
D004792:Enzyme Precursors; D021122:Protein Subunits; D011994:Recombinant Proteins; D014357:Trypsin; D005341:Fibrinolysin; D008666:Metalloendopeptidases; C053833:meprin A; C095663:meprin B; D015032:Zinc