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Direct production of functional matrix metalloproteinase—14 without refolding or activation and its application for in vitro inhibition assays
Author(s) -
Nam Dong Hyun,
Ge Xin
Publication year - 2016
Publication title -
biotechnology and bioengineering
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.136
H-Index - 189
eISSN - 1097-0290
pISSN - 0006-3592
DOI - 10.1002/bit.25840
Subject(s) - periplasmic space , matrix metalloproteinase , chemistry , metalloproteinase , biochemistry , in vitro , biological activity , microbiology and biotechnology , escherichia coli , biology , gene
Human matrix metalloproteinase (MMP)‐14, a membrane‐bound zinc endopeptidase, is one of the most important cancer targets because it plays central roles in tumor growth and invasion. Large amounts of active MMP‐14 are required for cancer research and the development of chemical or biological MMP‐14 inhibitors. Current methods of MMP‐14 production through refolding and activation are labor‐intensive, time‐consuming, and often associated with low recovery rates, lot‐to‐lot variation and heterogeneous products. Here, we report direct production of the catalytic domain of MMP‐14 in the periplasmic space of Escherichia coli . 0.5 mg/L of functional MMP‐14 was produced without tedious refolding or problematic activation process. MMP‐14 prepared by simple periplasmic treatment can be readily utilized to evaluate the potencies of chemical and antibody‐based inhibitors. Furthermore, co‐expression of both MMP‐14 and antibody Fab fragments in the periplasm facilitated inhibitory antibody screening by avoiding purification of MMP‐14 or Fabs. We expect this MMP‐14 expression strategy can expedite the development of therapeutic drugs targeting MMPs with biological significance. Biotechnol. Bioeng. 2016;113: 717–723. © 2015 Wiley Periodicals, Inc.