Abstract
An antibacterial protein (about 12 kDa) was isolated from human amniotic fluid through dialysis, ultrafiltration and C18 reversed-phase HPLC steps. Automated Edman degradation showed that the N-terminal sequence of the antibacterial protein was NH(2)-Ile-Gln-Arg-Thr-Pro-Lys-Ile-Gln-Val-Tyr-Ser-Arg-His-Pro-Ala-Glu-Asn-Gly-. The N-terminal sequence of the antibacterial protein was found to be identical to that of β(2)-microglobulin, a component of MHC class I molecules, which are present on all nucleated cells. Matrix-assisted laser desorption ionization mass spectrometry (MALDI-MS) revealed that the molecular mass of the antibacterial protein was 11,631 Da. This antibacterial protein, β(2)M, possessed potent antibacterial activity against pathogenic bacteria. Specially, antibacterial activity was observed in potassium buffer, and potassium ion was found to be critical for the antibacterial activity. Interestingly, the antibacterial action of β(2)M was associated with dissipation of the transmembrane potential, but the protein did not cause damage to the membrane that would result in SYTOX green uptake. In addition, stimulation of WISH amniotic epithelial cells with the bacterial endotoxin lipopolysaccharide (LPS) induced dose-dependent upregulation of β(2)M mRNA expression. These results suggest that β(2)M contributes to a self-defense response when amniotic cells are exposed to pathogens.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Amino Acid Sequence
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Amniotic Fluid / cytology
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Amniotic Fluid / metabolism*
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Antimicrobial Cationic Peptides / genetics
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Antimicrobial Cationic Peptides / isolation & purification
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Antimicrobial Cationic Peptides / metabolism*
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Antimicrobial Cationic Peptides / pharmacology
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Cells, Cultured
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Chromatography, High Pressure Liquid
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Drug Resistance, Bacterial
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Epithelial Cells / immunology
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Epithelial Cells / metabolism
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Escherichia coli / drug effects
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Escherichia coli / physiology
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Female
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Gene Expression Regulation / immunology
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Humans
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Immunity, Innate
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Lipopolysaccharides / pharmacology
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Listeria monocytogenes / drug effects
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Listeria monocytogenes / physiology
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Membrane Potentials / drug effects
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Microbial Sensitivity Tests
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Molecular Sequence Data
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Pregnancy
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RNA, Messenger / genetics
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RNA, Messenger / metabolism
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Sequence Analysis, Protein
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beta 2-Microglobulin / genetics
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beta 2-Microglobulin / isolation & purification
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beta 2-Microglobulin / metabolism*
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beta 2-Microglobulin / pharmacology
Substances
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Antimicrobial Cationic Peptides
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Lipopolysaccharides
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RNA, Messenger
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beta 2-Microglobulin
Grants and funding
This work was supported by the National Research Foundation of Korea grant funded by the Korea government (MEST) (No. 2011-0017532). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.