Journal
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
Volume 112, Issue 37, Pages 11660-11665Publisher
NATL ACAD SCIENCES
DOI: 10.1073/pnas.1514026112
Keywords
chlamydia; peptidoglycan; mass spectrometry; NOD2 receptor; muropeptide
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Funding
- National Institute of Allergy and Infectious Diseases [R56 AI044033]
- NIH Centers for AIDS Research at the Albert Einstein College of Medicine [AI-051519]
- NIH [AI26170]
- Bill and Melinda Gates Foundation [OPP1033104]
- Bill and Melinda Gates Foundation [OPP1033104] Funding Source: Bill and Melinda Gates Foundation
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The chlamydial anomaly, first coined by James Moulder, describes the inability of researchers to detect or purify peptidoglycan (PG) from pathogenic Chlamydiae despite genetic and biochemical evidence and antibiotic susceptibility data that suggest its existence. We recently detected PG in Chlamydia trachomatis by a new metabolic cell wall labeling method, however efforts to purify PG from pathogenic Chlamydiae have remained unsuccessful. Pathogenic chlamydial species are known to activate nucleotide-binding oligomerization domain-containing protein 2 (NOD2) innate immune receptors by as yet uncharacterized ligands, which are presumed to be PG fragments (muramyl di-and tripeptides). We used the NOD2-dependent activation of NF-kappa B by C. trachomatis-infected cell lysates as a biomarker for the presence of PG fragments within specific lysate fractions. We designed a new method of muropeptide isolation consisting of a double filtration step coupled with reverse-phase HPLC fractionation of Chlamydia-infected HeLa cell lysates. Fractions that displayed NOD2 activity were analyzed by electrospray ionization mass spectrometry, confirming the presence of muramyl di-and tripeptides in Chlamydia-infected cell lysate fractions. Moreover, the mass spectrometry data of large muropeptide fragments provided evidence that transpeptidation and transglycosylation reactions occur in pathogenic Chlamydiae. These results reveal the composition of chlamydial PG and disprove the glycanless peptidoglycan hypothesis.
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