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Published on: April 1, 2014
Binding of bacterial peptidoglycan to CD14
R Dziarski1, R I Tapping, P S Tobias
1Northwest Center for Medical Education, Indiana University School of Medicine, Gary, Indiana 46408, USA. rdziar@iunhaw1.iun.indiana.edu
The Journal of Biological Chemistry
|May 16, 1998
Summary
Soluble peptidoglycan (sPGN) activates macrophages by binding to CD14, a receptor typically for lipopolysaccharide (LPS). This study confirms sPGN-CD14 binding, revealing partially overlapping binding sites with LPS.
Area of Science:
- Immunology
- Microbiology
- Molecular Biology
Background:
- Soluble peptidoglycan (sPGN) from Gram-positive bacteria is a known macrophage activator.
- CD14 is a well-established receptor for lipopolysaccharide (LPS), a component of Gram-negative bacteria.
- The interaction between sPGN and CD14 has been hypothesized but required detailed investigation.
Purpose of the Study:
- To experimentally validate the hypothesis that soluble peptidoglycan (sPGN) binds to the CD14 receptor.
- To characterize the binding kinetics and specificity of sPGN interaction with CD14.
- To compare the binding of sPGN to CD14 with that of lipopolysaccharide (LPS).
Main Methods:
- Utilized three distinct binding assays: sPGN-immobilized assays with soluble 32P-labeled CD14 (sCD14), enzyme-linked immunosorbent assay (ELISA), and photoaffinity cross-linking.
- Investigated competitive inhibition using various molecules including unlabeled sCD14, CD14 fragments, LPS, lipid A, and lipoteichoic acid.
- Assessed the influence of LPS-binding protein (LBP) on sCD14 binding to sPGN and LPS.
- Employed anti-CD14 monoclonal antibodies (mAbs) to probe for distinct binding sites.
Main Results:
- Confirmed specific binding of sPGN to CD14 across all tested assays.
- Demonstrated that sPGN binds to CD14 with higher affinity (KD = 25 nM) than LPS (KD = 41 nM), although binding is slower.
- Showed that LPS-binding protein (LBP) modulates sCD14 binding differently for sPGN and LPS, enhancing affinity for LPS.
- Identified partially overlapping, yet distinct, conformational binding sites on CD14 for sPGN and LPS, evidenced by differential mAb inhibition.
Conclusions:
- Soluble peptidoglycan (sPGN) directly binds to CD14, extending the known ligand repertoire of this receptor beyond LPS.
- The binding interaction between sPGN and CD14 involves specific molecular recognition and exhibits distinct characteristics compared to LPS binding.
- These findings elucidate a novel mechanism for sPGN-mediated immune activation via CD14 and highlight the receptor's versatility.
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