IL-8/CXC ligand 8 survives neonatal gastric digestion as a result of intrinsic aspartyl proteinase resistance.
The human fetus and neonate swallow biologically significant quantities of IL-8/CXC ligand 8 (CXCL8) in amniotic fluid and breast milk, and this remains measurable through simulated neonatal gastric and proximal intestinal digestions. We sought to confirm the structural and functional integrity of IL-8/CXCL8 in digestates and determine the mechanisms underlying this protease resistance. We observed that in comparison with BSA, IL-8/CXCL8 is highly resistant to pepsin and can be detected intact in assays for structural, immunologic, and functional integrity. In a computational molecular docking simulation, IL-8/CXCL8 was observed to fit poorly in the pepsin active site. On the basis of simulated mutation analyses, we hypothesized that this protease resistance is due to disulfide bond-related tertiary folding in IL-8/CXCL8. This was confirmed on chemical reduction of these groups.
Maheshwari
Akhil
A
Division of Neonatology, Deparment of Pediatrics, University of South Florida College of Medicine and All Children's Hospital, St. Petersburg, FL 33701, USA. Akhil@peds.uab.edu
Lu
Wenge
W
Guida
Wayne C
WC
Christensen
Robert D
RD
Calhoun
Darlene A
DA
D000818:Animals; D001665:Binding Sites; D004063:Digestion; D004220:Disulfides / Q000737:chemistry; D004847:Epithelial Cells / Q000166:cytology / Q000378:metabolism; D005750:Gastric Juice / Q000201:enzymology; D005765:Gastrointestinal Agents / Q000378:metabolism; D006801:Humans; D007223:Infant; D007231:Infant, Newborn; D016209:Interleukin-8 / Q000737:chemistry* / Q000378:metabolism*; D008958:Models, Molecular; D009504:Neutrophils / Q000378:metabolism; D010434:Pepsin A / Q000378:metabolism*; D011487:Protein Conformation; D013270:Stomach / Q000166:cytology / Q000201:enzymology; D013552:Swine
D004220:Disulfides; D005765:Gastrointestinal Agents; D016209:Interleukin-8; D010434:Pepsin A