Purification from human milk of matriptase complexes with secreted serpins: mechanism for inhibition of matriptase other than HAI-1

Am J Physiol Cell Physiol. 2008 Aug;295(2):C423-31. doi: 10.1152/ajpcell.00164.2008. Epub 2008 Jun 11.

Abstract

Matriptase, a type 2 transmembrane serine protease, is predominately expressed by epithelial and carcinoma cells in which hepatocyte growth factor activator inhibitor 1 (HAI-1), a membrane-bound, Kunitz-type serine protease inhibitor, is also expressed. HAI-1 plays dual roles in the regulation of matriptase, as a conventional protease inhibitor and as a factor required for zymogen activation of matriptase. As a consequence, activation of matriptase is immediately followed by HAI-1-mediated inhibition, with the activated matriptase being sequestered into HAI-1 complexes. Matriptase is also expressed by peripheral blood leukocytes, such as monocytes and macrophages; however, in contrast to epithelial cells, monocytes and macrophages were reported not to express HAI-1, suggesting that these leukocytes possess alternate, HAI-1-independent mechanisms regulating the zymogen activation and protease inhibition of matriptase. In the present study, we characterized matriptase complexes of 110 kDa in human milk, which contained no HAI-1 and resisted dissociation in boiling SDS in the absence of reducing agents. These complexes were further purified and dissociated into 80-kDa and 45-kDa fragments by treatment with reducing agents. Proteomic and immunological methods identified the 45-kDa fragment as the noncatalytic domains of matriptase and the 80-kDa fragment as the matriptase serine protease domain covalently linked to one of three different secreted serpin inhibitors: antithrombin III, alpha1-antitrypsin, and alpha2-antiplasmin. Identification of matriptase-serpin inhibitor complexes provides evidence for the first time that the proteolytic activity of matriptase, from those cells that express no or low levels of HAI-1, may be controlled by secreted serpins.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Antithrombin III / analysis
  • Antithrombin III / isolation & purification
  • Antithrombin III / metabolism
  • Blotting, Western
  • Chromatography, Liquid
  • Electrophoresis, Polyacrylamide Gel
  • Humans
  • Mass Spectrometry
  • Milk, Human / chemistry
  • Milk, Human / metabolism*
  • Molecular Sequence Data
  • Proteinase Inhibitory Proteins, Secretory / analysis
  • Proteinase Inhibitory Proteins, Secretory / isolation & purification
  • Proteinase Inhibitory Proteins, Secretory / metabolism
  • Serine Endopeptidases / analysis
  • Serine Endopeptidases / isolation & purification
  • Serine Endopeptidases / metabolism*
  • Serpins / analysis
  • Serpins / isolation & purification
  • Serpins / metabolism*
  • alpha 1-Antitrypsin / analysis
  • alpha 1-Antitrypsin / isolation & purification
  • alpha 1-Antitrypsin / metabolism
  • alpha-2-Antiplasmin / analysis
  • alpha-2-Antiplasmin / isolation & purification
  • alpha-2-Antiplasmin / metabolism

Substances

  • Proteinase Inhibitory Proteins, Secretory
  • SPINT1 protein, human
  • Serpins
  • alpha 1-Antitrypsin
  • alpha-2-Antiplasmin
  • Antithrombin III
  • Serine Endopeptidases
  • ST14 protein, human