Clusterin, an abundant serum factor, is a possible negative regulator of MT6-MMP/MMP-25 produced by neutrophils.
MT6-MMP/MMP-25 is the latest member of the membrane-type matrix metalloproteinase (MT-MMP) subgroup in the MMP family and is expressed in neutrophils and some brain tumors. The proteolytic activity of MT6-MMP has been studied using recombinant catalytic fragments and shown to degrade several components of the extracellular matrix. However, the activity is possibly modulated further by the C-terminal hemopexin-like domain, because some MMPs are known to interact with other proteins through this domain. To explore the possible function of this domain, we purified a recombinant MT6-MMP with the hemopexin-like domain as a soluble form using a Madin-Darby canine kidney cell line as a producer. Mature and soluble MT6-MMP processed at the furin motif was purified as a 45-kDa protein together with a 46-kDa protein having a single cleavage in the hemopexin-like domain. Interestingly, 73- and 70-kDa proteins were co-purified with the soluble MT6-MMP by forming stable complexes. They were identified as clusterin, a major component of serum, by N-terminal amino acid sequencing. MT1-MMP that also has a hemopexin-like domain did not form a complex with clusterin. MT6-MMP forming a complex with clusterin was detected in human neutrophils as well. The enzyme activity of the soluble MT6-MMP was inactive in the clusterin complex. Purified clusterin was inhibitory against the activity of soluble MT6-MMP. On the other hand, it had no effect on the activities of MMP-2 and soluble MT1-MMP. Because clusterin is an abundant protein in the body fluid in tissues, it may act as a negative regulator of MT6-MMP in vivo.
Matsuda
Akira
A
Division of Cancer Cell Research, Institute of Medical Science, the University of Tokyo, Minato-ku, Tokyo 108-8639, Japan.
Itoh
Yoshifumi
Y
Koshikawa
Naohiko
N
Akizawa
Toshifumi
T
Yana
Ikuo
I
Seiki
Motoharu
M
D020816:Amino Acid Motifs; D000818:Animals; D015153:Blotting, Western; D019556:COS Cells; D020134:Catalytic Domain; D002460:Cell Line; D045744:Cell Line, Tumor; D002850:Chromatography, Gel; D051152:Clusterin; D018076:DNA, Complementary / Q000378:metabolism; D004285:Dogs; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D000939:Epitopes; D045683:Furin / Q000737:chemistry; D058851:GPI-Linked Proteins; D005786:Gene Expression Regulation; D005822:Genetic Vectors; D006023:Glycoproteins / Q000737:chemistry* / Q000378:metabolism / Q000502:physiology*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D020782:Matrix Metalloproteinases / Q000096:biosynthesis* / Q000235:genetics; D053504:Matrix Metalloproteinases, Membrane-Associated; D008957:Models, Genetic; D018832:Molecular Chaperones / Q000737:chemistry* / Q000378:metabolism / Q000502:physiology*; D009504:Neutrophils / Q000378:metabolism*; D010957:Plasmids / Q000378:metabolism; D011233:Precipitin Tests; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D016622:Silver Staining; D013997:Time Factors; D014162:Transfection
C495242:CLU protein, human; D051152:Clusterin; D018076:DNA, Complementary; D000939:Epitopes; D058851:GPI-Linked Proteins; D006023:Glycoproteins; D018832:Molecular Chaperones; D011994:Recombinant Proteins; D045683:Furin; D020782:Matrix Metalloproteinases; D053504:Matrix Metalloproteinases, Membrane-Associated; C403928:matrix metalloproteinase 25