Engineering of tissue inhibitor of metalloproteinases TIMP-1 for fine discrimination between closely related stromelysins MMP-3 and MMP-10

J Biol Chem. 2022 Mar;298(3):101654. doi: 10.1016/j.jbc.2022.101654. Epub 2022 Jan 29.

Abstract

Matrix metalloproteinases (MMPs) have long been known as key drivers in the development and progression of diseases, including cancer and neurodegenerative, cardiovascular, and many other inflammatory and degenerative diseases, making them attractive potential drug targets. Engineering selective inhibitors based upon tissue inhibitors of metalloproteinases (TIMPs), endogenous human proteins that tightly yet nonspecifically bind to the family of MMPs, represents a promising new avenue for therapeutic development. Here, we used a counter-selective screening strategy for directed evolution of yeast-displayed human TIMP-1 to obtain TIMP-1 variants highly selective for the inhibition of MMP-3 in preference over MMP-10. As MMP-3 and MMP-10 are the most similar MMPs in sequence, structure, and function, our results thus clearly demonstrate the capability for engineering full-length TIMP proteins to be highly selective MMP inhibitors. We show using protein crystal structures and models of MMP-3-selective TIMP-1 variants bound to MMP-3 and counter-target MMP-10 how structural alterations within the N-terminal and C-terminal TIMP-1 domains create new favorable and selective interactions with MMP-3 and disrupt unique interactions with MMP-10. While our MMP-3-selective inhibitors may be of interest for future investigation in diseases where this enzyme drives pathology, our platform and screening strategy can be employed for developing selective inhibitors of additional MMPs implicated as therapeutic targets in disease.

Keywords: MMP; TIMP; crystal structure; directed evolution; matrix metalloproteinase; protease; protease inhibitor; protein engineering; tissue inhibitor of metalloproteinases; yeast surface display.

MeSH terms

  • Humans
  • Matrix Metalloproteinase 10 / chemistry
  • Matrix Metalloproteinase 10 / genetics
  • Matrix Metalloproteinase 10 / metabolism
  • Matrix Metalloproteinase 3* / chemistry
  • Matrix Metalloproteinase 3* / genetics
  • Matrix Metalloproteinase 3* / metabolism
  • Protein Engineering
  • Tissue Inhibitor of Metalloproteinase-1* / chemistry
  • Tissue Inhibitor of Metalloproteinase-1* / genetics
  • Tissue Inhibitor of Metalloproteinase-1* / metabolism

Substances

  • TIMP1 protein, human
  • Tissue Inhibitor of Metalloproteinase-1
  • MMP3 protein, human
  • Matrix Metalloproteinase 3
  • MMP10 protein, human
  • Matrix Metalloproteinase 10