Calprotectin inhibits matrix metalloproteinases by sequestration of zinc.
OBJECTIVE
Calprotectin, a 36 kDa protein present in neutrophil cytoplasm, has antimicrobial and apoptosis inducing activities, which are reversed by the addition of zinc. Matrix metalloproteinases (MMPs), a family of zinc dependent enzymes, are important in many normal biological processes including embryonic development, angiogenesis, and wound healing, but also pathological processes such as inflammation, cancer, and tissue destruction. The aim of this study was to investigate whether calprotectin can inhibit MMP activity, and whether such inhibition could be overcome by the addition of zinc.


METHODS
MMP activity was measured by the degradation of substrates precoated on to microwells, and visualised by Coomassie blue staining of residual substrate. Seven metalloproteinases (MMP-1, MMP-2, MMP-3, MMP-7, MMP-8, MMP-9, and MMP-13) were tested against two substrates: gelatin and alpha-casein.


RESULTS
All MMPs except MMP-1 were active against gelatin, whereas MMP-7 was the only enzyme active against alpha-casein. The addition of calprotectin inhibited the activity of all the MMPs, but different concentrations of the protein, from 0.3 microM to > 11microM, were necessary to produce a 50% inhibition of the MMPs. Inhibition by calprotectin was largely overcome by the addition of zinc.


CONCLUSIONS
The findings suggest that calprotectin inhibits MMPs by sequestration of zinc. The data also suggest that MMPs have different affinities for zinc and that calprotectin has a lower zinc affinity than the MMPs.
Isaksen
B
B
Department of Immunology and Transfusion Medicine, Ullevaal University Hospital, Kirkeveien 166, 0407 Oslo, Norway. barbro.isaksen@ulleval.no
Fagerhol
M K
MK
D002364:Caseins / Q000378:metabolism; D017364:Collagenases / Q000502:physiology; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005780:Gelatin / Q000378:metabolism; D006801:Humans; D039841:Leukocyte L1 Antigen Complex; D020781:Matrix Metalloproteinase 1 / Q000502:physiology; D053509:Matrix Metalloproteinase 13; D020778:Matrix Metalloproteinase 2 / Q000502:physiology; D019278:Matrix Metalloproteinase 3 / Q000502:physiology; D020783:Matrix Metalloproteinase 7 / Q000502:physiology; D020784:Matrix Metalloproteinase 8 / Q000502:physiology; D020780:Matrix Metalloproteinase 9 / Q000502:physiology; D061965:Matrix Metalloproteinase Inhibitors; D020782:Matrix Metalloproteinases / Q000502:physiology*; D008562:Membrane Glycoproteins / Q000502:physiology*; D019006:Neural Cell Adhesion Molecules / Q000502:physiology*; D015032:Zinc / Q000378:metabolism*
D002364:Caseins; D004791:Enzyme Inhibitors; D039841:Leukocyte L1 Antigen Complex; D061965:Matrix Metalloproteinase Inhibitors; D008562:Membrane Glycoproteins; D019006:Neural Cell Adhesion Molecules; D005780:Gelatin; D017364:Collagenases; C507079:MMP13 protein, human; D053509:Matrix Metalloproteinase 13; D020782:Matrix Metalloproteinases; D019278:Matrix Metalloproteinase 3; D020783:Matrix Metalloproteinase 7; D020778:Matrix Metalloproteinase 2; D020784:Matrix Metalloproteinase 8; D020780:Matrix Metalloproteinase 9; D020781:Matrix Metalloproteinase 1; D015032:Zinc