TY - JOUR
T1 - The serine protease motif of Pic mediates a dose-dependent mucolytic activity after binding to sugar constituents of the mucin substrate
AU - Gutiérrez-Jiménez, Javier
AU - Arciniega, Ivonne
AU - Navarro-García, Fernando
N1 - Funding Information:
We thank Ken Teter for the invaluable critical review of the manuscript and Lucia Chavez-Dueñas for technical help. This work was supported by grants from Consejo Nacional de Ciencia y Tecnología de México (CONACYT, 30004M and C02-44660) to FNG. JGJ was supported by a scholarship from Promep (SEP).
PY - 2008/8
Y1 - 2008/8
N2 - The pic gene is harbored on the chromosomes of three important pathogens: enteroaggregative Escherichia coli (EAEC), uropathogenic E. coli (UPEC), and Shigella flexneri. Since Pic is secreted into the intestinal lumen during EAEC infection, we sought to identify intestinal-mucosal substrates for Pic. Pic did not damage epithelial cells, cleave fodrin, or degrade host defense proteins embedded in the mucus layer (sIgA, lactoferrin and lysozyme). However, by using a solid-phase assay to evaluate the mucinolytic activity of EAEC Pic, we documented a specific, dose-dependent mucinolytic activity. A serine protease inhibitor and an enzymatically inactive variant of Pic were used to show that the Pic serine protease motif is required for mucinolytic activity. Pic binds mucin, and this binding was blocked in competition assays using monosaccharide constituents of the oligosaccharide side chains of mucin. Moreover, Pic mucinolytic activity decreased when sialic acid was removed from mucin. Thus, Pic is a mucinase with lectin-like activity that can be related to its reported hemagglutinin activity. Our results suggest that EAEC may secrete Pic into the intestinal lumen as a strategy for penetrating the gel-like mucus layer during EAEC colonization.
AB - The pic gene is harbored on the chromosomes of three important pathogens: enteroaggregative Escherichia coli (EAEC), uropathogenic E. coli (UPEC), and Shigella flexneri. Since Pic is secreted into the intestinal lumen during EAEC infection, we sought to identify intestinal-mucosal substrates for Pic. Pic did not damage epithelial cells, cleave fodrin, or degrade host defense proteins embedded in the mucus layer (sIgA, lactoferrin and lysozyme). However, by using a solid-phase assay to evaluate the mucinolytic activity of EAEC Pic, we documented a specific, dose-dependent mucinolytic activity. A serine protease inhibitor and an enzymatically inactive variant of Pic were used to show that the Pic serine protease motif is required for mucinolytic activity. Pic binds mucin, and this binding was blocked in competition assays using monosaccharide constituents of the oligosaccharide side chains of mucin. Moreover, Pic mucinolytic activity decreased when sialic acid was removed from mucin. Thus, Pic is a mucinase with lectin-like activity that can be related to its reported hemagglutinin activity. Our results suggest that EAEC may secrete Pic into the intestinal lumen as a strategy for penetrating the gel-like mucus layer during EAEC colonization.
KW - Autotransporter protein
KW - Lectin activity
KW - Mucin
KW - Mucinase
KW - Pic
KW - Proteolytic activity
UR - http://www.scopus.com/inward/record.url?scp=47049125726&partnerID=8YFLogxK
U2 - 10.1016/j.micpath.2008.04.006
DO - 10.1016/j.micpath.2008.04.006
M3 - Artículo
C2 - 18538533
AN - SCOPUS:47049125726
SN - 0882-4010
VL - 45
SP - 115
EP - 123
JO - Microbial Pathogenesis
JF - Microbial Pathogenesis
IS - 2
ER -