Interaction of human salivary mucin MG2, its recombinant N-terminal region and a synthetic peptide with Actinobacillus actinomycetemcomitans.
The antimicrobial properties of human salivary mucin MG2 against the periodontal pathogen, Actinobacillus actinomycetemcomitans (A. actinomycetemcomitans), were investigated using purified MG2, rNMUC7 (a recombinant polypeptide containing residue 1-144 of MG2) and synthetic peptides PEP1 (residue 1-17) and PEP2 (residue 47-63). MG2 and rNMUC7 bound to A. actinomycetemcomitans strains SUNY75, SUNY465, SUNY523, 652 and JP2 in a liquid phase binding assay. The bactericidal activities of rNMUC7, PEP1 and PEP2 against A. actinomycetemcomitans SUNY523 were examined in a colony forming unit killing assay. The LD50 for rNMUC7 was 9 microM, for PEP2 was 20 microM and PEP1 did not exhibit bactericidal activity. The primary structure of these polypeptides was analyzed and a direct relationship between net positive charge and bactericidal activity was found. Screening of saliva samples from 60 individuals on Western blots probed with an anti-MG2 antibody against PEP2 revealed that a 20 kDa MG2 fragment was present in 66% of subjects and that this fragment was not present in glandular secretions. Matrix-assisted laser desorption ionization time-of-flight mass spectrometry of tryptic peptides derived from the 20 kDa fragment confirmed that this fragment contained a portion of the amino terminal region of MG2. The present study showed that the N-terminal region of MG2 and a subdomain within this region are microbicidal against A. actinomycetemcomitans and that a 20 kDa fragment of MG2 occurs in whole saliva. This suggests that cleavage of MG2 in vivo may produce fragments with microbicidal properties and that this may represent a novel mechanism of host defense.
Liu
Bing
B
Department of Biochemistry, Boston University School of Medicine, Boston, MA 02118, USA.
Rayment
Sean A
SA
Soares
Rodrigo V
RV
Oppenheim
Frank G
FG
Offner
Gwynneth D
GD
Fives-Taylor
Paula
P
Troxler
Robert F
RF
D000328:Adult; D016976:Aggregatibacter actinomycetemcomitans / Q000187:drug effects* / Q000254:growth & development; D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D015153:Blotting, Western; D015169:Colony Count, Microbial; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D007525:Isoelectric Focusing; D013058:Mass Spectrometry; D008875:Middle Aged; D009077:Mucins / Q000138:chemical synthesis / Q000494:pharmacology*; D011485:Protein Binding; D011994:Recombinant Proteins; D012463:Saliva / Q000737:chemistry; D012471:Salivary Proteins and Peptides / Q000138:chemical synthesis / Q000494:pharmacology*
D000900:Anti-Bacterial Agents; C083238:MUC7 protein, human; D009077:Mucins; D011994:Recombinant Proteins; D012471:Salivary Proteins and Peptides