Glycosylation broadens the substrate profile of membrane type 1 matrix metalloproteinase.

Membrane type 1 matrix metalloproteinase (MT1-MMP) is a collagenolytic enzyme that has been implicated in normal development and in pathological processes such as cancer metastasis. The activity of MT1-MMP is regulated extensively at the post-translational level, and the current data support the hypothesis that MT1-MMP activity is modulated by glycosylation. Enzymatic deglycosylation, site-directed mutagenesis, and lectin precipitation assays were used to demonstrate that MT1-MMP contains O-linked complex carbohydrates on the Thr(291), Thr(299), Thr(300), and/or Ser(301) residues in the proline-rich linker region. MT1-MMP glycoforms were detected in human cancer cell lines, suggesting that MT1-MMP activity may be regulated by differential glycosylation in vivo. Although the autolytic processing and interstitial collagenase activity of MT1-MMP were not impaired in glycosylation-deficient mutants, cell surface MT1-MMP-catalyzed activation of pro-matrix metalloproteinase-2 (proMMP-2) required proper glycosylation of MT1-MMP. The inability of carbohydrate-free MT1-MMP to activate proMMP-2 was not a result of defective MT1-MMP zymogen activation, aberrant protein stability, or inability of the mature enzyme to oligomerize. Rather, our data support a mechanism whereby glycosylation affects the recruitment of tissue inhibitor of metalloproteinases-2 (TIMP-2) to the cell surface, resulting in defective formation of the MT1-MMP/TIMP-2/proMMP-2 trimeric activation complex. These data provide evidence for an additional mechanism for post-translational control of MT1-MMP activity and suggest that glycosylation of MT1-MMP may regulate its substrate targeting.